Aromatic heterocyclic derivatives as enzyme inhibitors

ABSTRACT

The present invention discloses peptide aldehydes which are potent and specific inhibitors of thrombin, their pharmaceutically acceptable salts, pharmaceutically acceptable compositions thereof, and methods of using them as therapeutic agents for disease states in mammals characterized by abnormal thrombosis.

CROSS REFERENCE TO RELATED APPLICATION

This application is a continuation-in-part of U.S. Ser. No. 08/573,775,filed Dec. 18, 1995 which is a continuation-in-part of U.S. Ser. No.08/481,660 now U.S. Pat. No. 5,658,930 and U.S. Ser. No. 08/484,506, nowU.S. Pat. No. 5,656,645 both filed Jun. 7, 1995 and both of which arecontinuations-in-part of U.S. Ser. No. 08/356,833, filed Dec. 13, 1994;the disclosures of all these applications are incorporated herein byreference.

TECHNICAL FIELDS

In one aspect, the present invention relates to compounds which arepotent and specific inhibitors of thrombin. In another aspect, thepresent invention relates to novel peptide aldehydes, theirpharmaceutically acceptable salts, and pharmaceutically acceptablecompositions thereof which are useful as potent and specific inhibitorsof blood coagulation in vitro and in vivo in mammals. In yet anotheraspect, the invention relates to methods of using these inhibitors astherapeutic agents for disease states in mammals characterized byabnormal thrombosis.

BACKGROUND

Normal hemostasis is the result of a complex balance between theprocesses of clot formation (blood coagulation) and clot dissolution(fibrinolysis). The complex interactions between blood cells, specificplasma proteins and the vascular surface, maintain the fluidity of bloodunless injury and blood loss occur.

Blood coagulation is the culmination of a series of amplified reactionsin which several specific zymogens of serine proteases in plasma areactivated by limited proteolysis. Nemerson, Y. and Nossel, H. L., Ann.Rev. Med., 33:479 (1982). This series of reactions results in theformation of an insoluble fibrin matrix which is required for thestabilization of the primary hemostatic plug. The interaction andpropagation of the activation reactions occurs through the extrinsic andintrinsic pathways of coagulation.

These pathways are highly inter-dependent and converge in the formationof the serine protease, Factor Xa. Factor Xa catalyzes the penultimatestep in the blood coagulation cascade which is the formation of theserine protease thrombin. This step occurs following the assembly of theprothrombinase complex which is composed of factor Xa, the non-enzymaticco-factor Va and the substrate prothrombin assembled on the surface ofadhered, activated platelets or systemically circulating membranousmicroparticles.

Proteolytic activation of zymogen factor X to its catalytically activeform, factor Xa, can occur by either the intrinsic or extrinsiccoagulation pathways.

The intrinsic pathway is referred to as "intrinsic" because everythingneeded for clotting is in the blood. Saito, H., "Normal HemostaticMechanisms", Disorders of Hemostasis, pp. 27-29, Grune & Stratton, Inc.(O. D. Ratnoff, M.D. and C. D. Forbes, M.D. edit. 1984). This pathway iscomprised of the zymogen serine proteases, factors IX and XI, and thenon-enzymatic co-factor, factor VIII. The initiation of the intrinsicpathway results in the activation of factor XI to XIa. Factor XIacatalyzes the activation of factor IX to factor IXa which in combinationwith the activated form of factor VIII on an appropriate phospholipidsurface, results in the formation of the tenase complex. This complexalso catalyzes the formation of the serine protease, factor Xa, from itszymogen, factor X which subsequently results in clot formation.

The extrinsic pathway is referred to as "extrinsic" because the tissuefactor which binds to and facilitates the activation of factor VII comesfrom outside the blood. Saito, Id. The major components of this pathwayare the zymogen serine protease, factor VII, and the membrane boundprotein, tissue factor. The latter serves as the requisite non-enzymaticco-factor for this enzyme. The initiation of this pathway is thought tobe an autocatalytic event resulting from the activation of zymogenfactor VII by trace levels of activated factor VII (factor VIIa), bothof which are bound to newly exposed tissue factor on membrane surfacesat sites of vascular damage. The factor VIIa/tissue factor complexdirectly catalyzes the formation of the serine protease, factor Xa, fromits zymogen, factor X. Exposure of blood to injured tissue initiatesblood clotting by the extrinsic pathway.

The formation of thrombin is catalyzed by factor Xa following theassembly of the catalytic prothrombinase complex as reviewed by Mann, K.G. et al., "Surface-Dependent Reactions of the Vitamin K-DependentEnzyme Complexes", Blood, 76:1-16 (1990). This complex is composed offactor Xa, the non-enzymatic co-factor Va and the substrate prothrombinall assembled on an appropriate phospholipid surface. The requirement ofa macromolecular complex for efficient catalysis results in theprotection of factor Xa from natural anticoagulant mechanisms such asheparin-antithrombin III mediated inhibition. Teite, J. M. andRosenberg, R. D., "Protection of Factor Xa from neutralization by theheparin-antithrombin complex", J. Clin. Invest., 71:1383-1391(1983). Inaddition, sequestration of factor Xa in the prothrombinase complex alsorenders it resistant to inhibition by exogenous heparin therapy whichalso requires antithrombin III to elicit its anticoagulant effect.

Thrombin is the primary mediator of thrombus formation. Thrombin actsdirectly to cause formation of insoluble fibrin from circulatingfibrinogen. In addition, thrombin activates the zymogen factor XIII tothe active transglutaminase factor XIIIa which acts to covalentlystabilize the growing thrombus by crosslinking the fibrin strands.Lorand, L. and Konishi, K., Arch. Biochem. Biophys., 105:58 (1964).Beyond its direct role in the formation and stabilization of fibrin richclots, the enzyme has been reported to have profound bioregulatoryeffects on a number of cellular components within the vasculature andblood. Shuman, M. A., Ann. NY Acad. Sci., 405:349 (1986).

It is believed that thrombin is the most potent agonist of plateletactivation, and it has been demonstrated to be the primarypathophysiologic-mediator of platelet-dependent arterial thrombusformation. Edit, J. F. et al., J. Clin. Invest., 84:18 (1989).Thrombin-mediated platelet activation leads to ligand-inducedinter-platelet aggregation principally due to the bivalent interactionsbetween adhesive ligands such as fibrinogen and fibronectin withplatelet integrin receptors such as glycoprotein IIb/IIIa which assumetheir active conformation following thrombin activation. Berndt, M. C.and Phillips, D. R., Platelets in Biology and Pathology, pp 43-74,Elsevier/North Holland Biomedical Press (Gordon, J. L. edit. 1981).Thrombin-activated platelets can also support further thrombinproduction through the assembly of new prothrombinase and tenase (factorIXa, factor VIIIa and factor X) catalytic complexes on the membranesurface of intact activated platelets and platelet-derivedmicroparticles, following thrombin-mediated activation of thenon-enzymatic cofactors V and VIII, respectively. Tans, G. et al.,Blood, 77:2641 (1991). This positive feedback process results in thelocal generation of large concentrations of thrombin within the vicinityof the thrombus which supports further thrombus growth and extension.Mann, K. G. et al., Blood, 76:1 (1990).

In contrast to its prothrombotic effects, thrombin has been shown toinfluence other aspects of hemostasis. These include its effect as animportant physiological anticoagulant. The anticoagulant effect ofthrombin is expressed following binding of thrombin to the endothelialcell membrane glycoprotein, thrombomodulin. This is thought to result inan alteration of the substrate specificity of thrombin thereby allowingit to recognize and proteolytically activate circulating protein C togive activated protein C (aPC). Musci, G. et al., Biochemistry, 27:769(1988). aPC is a serine protease which selectively inactivates thenon-enzymatic co-factors Va and VIIIa resulting in a down-regulation ofthrombin formation by the prothrombinase and tenase catalytic complexes,respectively. Esmon, C. T., Science, 235:1348 (1987). The activation ofprotein C by thrombin in the absence of thrombomodulin is poor.

Thrombin has also been shown to be a potent direct mitogen for a numberof cell types, including cells of mesenchymal origin such as vascularsmooth muscle cells. Chen, L. B. and Buchanan, J. M., Proc. Natl. Acad.Sci. USA, 72:131 (1975). The direct interaction of thrombin withvascular smooth muscle also results in vasoconstriction. Walz, D. A. etal., Proc. Soc. Expl. Biol. Med., 180:518 (1985). Thrombin acts as adirect secretagogue inducing the release of a number of bioactivesubstances from vascular endothelial cells including tissue plasminogenactivator. Levin, E. G. et al., Thromb. Haemost., 56:115 (1986). Inaddition to these direct effects on vascular cells, the enzyme canindirectly elaborate potent mitogenic activity on vascular smooth musclecells by the release of several potent growth factors (e.g.,platelet-derived growth factor and epidermal growth factor) fromplatelet a-granules following thrombin-induced activation. Ross, R., N.Engl. J. Med., 314:408 (1986).

Many significant disease states are related to abnormal hemostasis. Withrespect to the coronary arterial vasculature, abnormal thrombusformation due to the rupture of an established atherosclerotic plaque isthe major cause of acute myocardial infarction and unstable angina.Moreover, treatment of an occlusive coronary thrombus by eitherthrombolytic therapy or percutaneous transluminal coronary angioplasty(PTCA) is often accompanied by an acute thrombotic reclosure of theaffected vessel which requires immediate resolution. With respect to thevenous vasculature, a high percentage of patients undergoing majorsurgery in the lower extremities or the abdominal area suffer fromthrombus formation in the venous vasculature which can result in reducedblood flow to the affected extremity and a predisposition to pulmonaryembolism. Disseminated intravascular coagulopathy commonly occurs withinboth vascular systems during septic shock, certain viral infections andcancer and is characterized by the rapid consumption of coagulationfactors and systemic coagulation which results in the formation oflife-threatening thrombi occurring throughout the vasculature leading towidespread organ failure.

Pathogenic thrombosis in the arterial vasculature is a major clinicalconcern in today's medicine. It is the leading cause of acute myocardialinfarction which is one of the leading causes of death in the westernworld. Recurrent arterial thrombosis also remains one of the leadingcauses of failure following enzymatic or mechanical recanalization ofoccluded coronary vessels using thrombolytic agents or percutaneoustransluminal coronary angioplasty (PTCA), respectively. Ross, A. M.,Thrombosis in Cardiovascular Disorder, p. 327, W.B. Saunders Co.(Fuster, V. and Verstraete, M. edit. 1991); Califf, R. M. and Willerson,J. T., Id. at p 389. In contrast to thrombotic events in the venousvasculature, arterial thrombosis is the result of a complex interactionbetween fibrin formation resulting from the blood coagulation cascadeand cellular components, particularly platelets, which make up a largepercentage of arterial thrombi. Heparin, the most widely used clinicalanticoagulant administered i.v., has not been shown to be universallyeffective in the treatment or prevention of acute arterial thrombosis orrethrombosis. Prins, M. H. and Hirsh, J., J. Am. Coll. Cardiol., 67:3A(1991).

Besides the unpredictable, recurrent thrombotic reocclusion whichcommonly occurs following PTCA, a profound restenosis of the recanalizedvessel occurs in 30 to 40% of patients 1 to 6 months following thisprocedure. Califf, R. M. et al., J. Am. Coll. Cardiol., 17:2B (1991).These patients require further treatment with either a repeat PTCA orcoronary artery bypass surgery to relieve the newly formed stenosis.Restenosis of a mechanically damaged vessel is not a thrombotic processbut instead is the result of a hyperproliferative response in thesurrounding smooth muscle cells which over time results in a decreasedluminal diameter of the affected vessel due to increased muscle mass.Id. As for arterial thrombosis, there is currently no effectivepharmacologic treatment for the prevention of vascular restenosisfollowing mechanical recanalization.

The need for safe and effective therapeutic anticoagulants has in oneaspect focused on the role of the serine protease thrombin in bloodcoagulation.

Most preferred natural substrates for thrombin are reported to containan uncharged amino acid in the P3 recognition subsite. For example, thethrombin cleavage site on the Aa chain of fibrinogen, which is theprimary physiological substrate for thrombin, is reported to contain aglycine residue in this position while the cleavage site on the Bb chaincontains a serine, as shown below:

P4 P3 P2 P1 P1'

Gly-Gly-Val-Arg/Gly Fibrinogen Aa Chain

Phe-Ser-Ala-Arg/Gly Fibrinogen Bb Chain

Peptidyl derivatives having an uncharged residue in the P3 position aresaid to bind to the active site of thrombin and thereby inhibit theconversion of fibrinogen to fibrin and inhibit cellular activation.These derivatives have either an aldehyde, chloromethyl ketone orboronic acid functionality associated with the P1 amino acid. Forexample, substrate-like peptidyl derivatives such asD-phenylalanyl-prolyl-argininal (D-Phe-Pro-Arg-al),D-phenylalanyl-prolyl-arginine-chloromethyl ketone (P-PACK) andacetyl-D-phenylalanyl-prolyl-boroarginine (Ac-(D-Phe)-Pro-boroArg) havebeen reported to inhibit thrombin by directly binding to the active siteof the enzyme. Bajusz, S., Symposia Biologica Hungarica, 25:277 (1984),Bajusz, S. et al, J. Med. Chem., 33:1729 (1990) and Bajusz, S. et al.,Int. J. Peptide Protein Res. 12:217 (1970); Kettner, C. and Shaw, E.,Methods Enzymol., 80:826 (1987), Kettner, C. et al., EP 293,881(published Dec. 7, 1988), Kettner, C., et al., J. Biol. Chem., 265:18209(1990). These molecules have been reported to be potent anticoagulantsin the prevention of platelet-rich arterial thrombosis. Kelly, A. B. etal., Thromb. Haemostas., 65:736 at abstract 257 (1991). Other peptidylaldehydes have been proposed or reported as inhibitors of thrombin. Bey,P. et al., EP 363,284 (published Apr. 11, 1990) and Balasubramanian, N.et al., EP 526,877 (published Feb. 10, 1993).

Peptidyl compounds which are said to be active site inhibitors ofthrombin but which differ in structure from those containing a unchargedamino acid in the P3 recognition subsite have been reported.

The compound, Argatroban (also called2R,4R-4-methyl-1-[N-2-(3-methyl-1,2,3,4-tetrahydro-8-quinolinesulfonyl)-L-argininal]-2-piperdinecarboxylicacid), is also reported to bind directly to the active site of thrombinand has been thought to be the most potent and selective compound in theclass of non-peptidyl inhibitors of this enzyme. Okamoto, S. et al.,Biochem. Biophys. Res. Commun., 101:440 (1981). Argatroban has beenreported to be a potent antithrombotic agent in several experimentalmodels of acute arterial thrombosis. Jang, I. K. et al., in bothCirculation, 81:219 (1990) and Circ. Res., 67:1552 (1990).

Peptidyl compounds which are said to be inhibitors of thrombin and whosemode of action is thought to be by binding to both the active site andanother site on the enzyme have been reported. Hirudin and certainpeptidyl derivatives of hirudin have been reported to inhibit bothconversion of fibrinogen to fibrin and platelet activation by binding toeither both the active site and exo site, or the exo site only, ofthrombin. Markwardt, F., Thromb. Haemostas., 66:141 (1991). Hirudin isreported to be a 65 amino acid polypeptide originally isolated fromleech salivary gland extracts. It is said to be one of the most potentinhibitors of thrombin known. Marki, W. E. and Wallis, R. B., Thromb.Haemostas., 64:344 (1990). It has been reported to inhibit thrombin bybinding to both its anion-binding exo-site and to its catalytic activesite which are distinct and physically distant from each other. Rydel,T. J. et al., Science, 249:277 (1990). Hirudin has been reported to be apotent antithrombotic agent in vivo. Markwardt, F. et al., Pharmazie,43:202 (1988); Kelly, A. B. et al., Blood, 77:1 (1991). In addition toits antithrombotic effects, hirudin has been reported to alsoeffectively inhibit smooth muscle proliferation and the associatedrestenosis following mechanical damage to a atherosclerotic rabbitfemoral artery. Sarembock, I. J. et al., Circulation, 84:232 (1991).

Hirugen has been reported to be a peptide derived from the anioniccarboxy-terminus of hirudin. It is reported to bind only to the anionbinding exo-site of thrombin and thereby inhibit the formation of fibrinbut not the catalytic turnover of small synthetic substrates which haveaccess to the unblocked active site of the enzyme. Maraganore, J. M. etal., J. Biol. Chem., 264:8692 (1989); Naski, M. C. et al., J. Biol.Chem., 265:13484 (1990). Based on x-ray crystallographic analysis, ithas been reported that the region of hirudin represented by hirugenbinds directly to the exo site of thrombin. Skrzypczak-Jankun, E. etal., Thromb. Haemostas., 65:830 at abstract 507 (1991). Moreover, thebinding of hirugen has also been reported to enhance the catalyticturnover of certain small synthetic substrates by thrombin, indicatingthat a conformational change in the enzyme active site may accompanyoccupancy of the exo-site. Liu, L. W. et al., J. Biol. Chem, 266:16977(1991). Hirugen also is reported to block thrombin-mediated plateletaggregation. Jakubowski, J. A. and Maraganore, J. M., Blood, 75:399(1990).

A group of synthetic chimeric molecules comprised of a hirugen-likesequence linked by a glycine-spacer region to the peptide,D-phenylalanyl-prolyl-arginine, which is based on a preferred substraterecognition site for thrombin, has been termed to be hirulog. Maraganoreet al., U.S. Pat. No. 5,196,404 (Mar. 23, 1993). The hirugen-likesequence is said to be linked to this peptide through the C-terminal endof the peptide. Maraganone, J. M. et al., Biochemistry, 29:7095 (1990).The hirulogs have been reported to be an effective antithrombotic agentsin preventing both fibrin-rich and platelet-rich thrombosis. Maraganone,J. M. et al., Thromb. Haemostas., 65:651 at abstract 17 (1991).

Certain benzamidines have been reported to inhibit thrombin thoughnon-selectively. 4-amidinophenylpyruvic acid (APPA) has been reported tobe a thrombin inhibitor with low toxicity and favorablepharmacokinetics. However, this compound was reported to benon-selective, inhibiting trypsin, plasmin and kallikrein. Markwardt etal., Thromb. Res., 1:243-52 (1972). Other benzamidine-derived structureswhich have been reported to inhibit thrombin include the cylic amides ofN^(a) -substituted 4-amidinophenylalanine and2-amino-5-(4-amidinophenyl)-1-valeric acid. The inhibitory constantdisplayed by these compounds was reported to be in the micromolar range.Markwardt et al., Thromb. Res., 17:425-31 (1980). Moreover, derivativesof 4-amidinophenylalanine whose a-amino group is linked to thearylsulfonyl residue via an w-aminoalkylcarboxylic acid as spacer havealso been assessed for their inhibitory effect. Among these N^(a)-(2-naphthylsulphonylglycyl)-4-amidino-phenylalanine piperidide(a-NAPAP) has been reported to possess an affinity for thrombin (K_(i)=6×10⁻⁹ M). Banner et al., J. Biol. Chem., 266:20085 (1991) andSturzebecher et al., Thromb. Res., 29:635-42 (1983).

Certain bis-benzamidines have been reported to inhibit thrombin. Theantithrombin activity of bis-benzamidines was reported to increase withthe length and bulkiness of the central chain. However, these compoundswere reported to be generally toxic in the micromolar range where theyare also inhibitory. Geratz et al., Thromb. Diath. Haemorrh., 29:154-67(1973); Geratz et al., J. Med. Chem., 16:970-5 (1973); Geratz et al., J.Med. Chem., 19:634-9 (1976); Walsmann et al., Acta Biol. Med. Germ.,35:K1-8 (1976); and Hauptmann et al., Acta Biol. Med. Germ., 35:635-44(1976).

Certain amidino-bearing aromatic ring structures such asbeta-naphthamidines have been reported to possess modest antithrombinand anticoagulant activity. This class of compounds include thenon-selective 6-amidino-2-naphthyl-4-guanidinobenzoatedimethanesulfonate (FUT 175). Fuji et al., Biochim. Biophys. Acta,661:342-5 (1981); and Hitomi et al., Haemostasis, 15:164-8 (1985).

Certain phenylguanidines have been reported to inhibit thrombin.Derivatives of 4-guanidinophenylalanine with inhibitory constants in themicromolar range have been reported to inhibit thrombin. This classincludes the N^(a) -tosylated and dansylated 4-guanidino phenylalaninepiperidides. Claeson et al., Thromb. Haemostas., 50:53 (1983). Anothercompound, [ethyl p-(6-guanidinohexanoyloxy) benzoate] methane sulfonate(FOY) was reported to be a non-selective competitive inhibitor ofthrombin. Ohno et al., Thromb. Res., 19:579-588 (1980).

SUMMARY OF THE INVENTION

The present invention is directed to novel peptide aldehyde compoundshaving arginine or arginine mimics at P₁ and pyridone, pyrimidone, oruracil groups as part of the peptide backbone. These compounds arepotent inhibitors of thrombin in vivo and in vitro.

Thus, in one aspect, the present invention is directed to compounds ofthe formula: ##STR1## wherein

(a) X is selected from the group consisting of --S(O)₂ --,--N(R')--S(O)₂ --, --(C═O)--, --OC(═O)--, --NH--C(═O)--, --P(O) (R")--and a direct link, wherein R' is hydrogen, alkyl of 1 to about 4 carbonatoms, aryl of about 6 to about 14 carbon atoms or aralkyl of about 6 toabout 16 carbon atoms, and R" is NR', OR', R', or SR', with the provisothat R" is not NH, OH, H, or SH, and;

(b) R₁ is selected from the group consisting of:

(1) alkyl of 1 to about 12 carbon atoms,

(2) alkyl of 1 to about 3 carbon atoms substituted with cyclic alkyl ofabout 3 to about 8 carbon atoms, which optionally is substituted in thering carbons with hydroxyl, amino, guanidino, amidino, or alkoxyl oralkyl each of 1 to about 3 carbons,

(3) cyclic alkyl of 3 to about 15 carbon atoms, which optionally issubstituted in the ring carbons with hydroxyl, amino, guanidino,amidino, or alkoxyl or alkyl each of 1 to about 3 carbons,

(4) heterocycloalkyl of 4 to about 10 ring atoms with the ring atomsselected from carbon and heteroatoms, wherein the heteroatoms areselected from the group consisting of oxygen, nitrogen, and S(O)i,wherein i is 0, 1 or 2, and which is optionally substituted on the ringcarbons with hydroxyl, alkoxyl or alkyl each of 1 to about 3 carbons,amino, guanidino, or amidino,

(5) heterocyclo of 4 to about 10 ring atoms with the ring atoms selectedfrom carbon and heteroatoms, wherein the heteroatoms are selected fromthe group consisting of oxygen, nitrogen, and S(O)_(i), wherein i is 0,1 or 2, including the group ##STR2## wherein ##STR3## is a 5 to 7 memberheterocycle of 3 to 6 ring carbon atoms, where V is --CH₂ --, --O--,--S(═O)--, --S(O)₂ -- or --S--, and which is optionally substituted onthe ring carbons with hydroxyl, alkoxyl or alkyl each of 1 to about 3carbons, amino, guanidino, or amidino,

(6) alkenyl of 2 to about 6 carbon atoms which is optionally substitutedwith cyclic alkyl of about 3 to about 8 carbon atoms, which optionallyis substituted in the ring carbons with hydroxyl, amino, guanidino,amidino, or alkoxyl or alkyl of 1 to about 3 carbons,

(7) aryl of about 6 to about 14 carbon atoms which is optionally mono-,di- or tri-substituted with Y₁, Y₂, and/or Y₃,

(8) heteroaryl of 5 to 14 atoms with the ring atoms selected from carbonand heteroatoms, wherein the heteroatoms are selected from oxygen,nitrogen, and S(O)_(i), wherein i is 0, 1 or 2, and which is optionallymono-, di- or tri-substituted with Y₁, Y₂, and/or Y₃,

(9) aralkyl of about 7 to about 15 carbon atoms which is optionallysubstituted on the alkyl chain with hydroxy or halogen and optionallymono-, di-, or tri-substituted on the aryl ring with Y₁, Y₂, and/or Y₃,

(10) heteroaralkyl of 6 to 10 atoms with the ring atoms selected fromcarbon and heteroatoms, wherein the heteroatoms are selected fromoxygen, nitrogen, and S(O)_(i), wherein i is 0, 1 or 2, and which isoptionally substituted on the alkyl chain with hydroxy or halogen andoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂, and/orY₃,

(11) aralkenyl of about 8 to about 15 carbon atoms which is optionallymono-, di-, or tri-substituted on the aryl ring with Y₁, Y₂, and/or Y₃,respectively,

(12) heteroaralkenyl of 7 to 12 atoms with the ring atoms selected fromcarbon and heteroatoms, wherein the heteroatoms are selected fromoxygen, nitrogen, and S(O)_(i), wherein i is 0, 1 or 2, and which isoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂, and/orY₃, ##STR4## (17) difluoromethyl and perfluoroalkyl of 1 to about 12carbon atoms, (18) perfluoroaryl of about 6 to about 14 carbon atoms,

(19) perfluoroaralkyl of about 7 to about 15 carbon atoms, and

(20) hydrogen,

wherein Y₁, Y₂, and Y₃ are

(i) independently selected from the group consisting of hydrogen,halogen, cyano, tetrazolyl, amino, guanidino, amidino, methylamino, andmethylguanidino, --CF₃, --CF₂ H, --CF₂ CF₃, --CH(CF₃)₂, --C(OH)(CF₃)₂,--OCF₃, --OCF₂ CF₃, --OC(O)NH₂, --OC(O)NHZ₁, --OC(O)NZ₁ Z₂, --NHC(O)Z₁,--NHC(O)NH₂, --NHC(O)NZ₁, --NHC(O)NZ₁ Z₂, --C(O)OH, --C(O)NH₂,--C(O)NHZ₁, --C(O)OZ₁, --P(O)₃ H, --P(O)₃ H₂, --P(O)₃ (Z₁)₂, --S(O)₃ H ,--S(O)_(m) Z₁, --Z₁, --OZ₁, --OH, --NH₂, --NHZ₁, and --NZ₁ Z₂, wherein mis 0, 1 or 2, and Z₁ and Z₂ are independently selected from the groupconsisting of alkyl of 1 to about 12 carbon atoms, aryl of about 6 toabout 14 carbon atoms, heteroaryl of about 5 to about 14 atoms having 1to about 9 carbon atoms, aralkyl of about 7 to about 15 carbon atoms,and heteroaralkyl of about 6 to about 11 atoms having about 3 to about 9carbon atoms, or

(ii) Y₁ and Y₂ are selected together to be --OC(Z₃)(Z₄)O--, wherein Z₃and Z₄ are independently selected from the group consisting of hydrogen,alkyl of 1 to about 12 carbon atoms, aryl of about 6 to about 14 carbonatoms, heteroaryl of about 5 to about 14 atoms having 1 to about 9carbon atoms, aralkyl of about 7 to about 15 carbon atoms, andheteroaralkyl of about 6 to about 11 atoms having about 3 to about 9carbon atoms, with the proviso that if X is not a direct link, then R₁is not hydrogen,

(c) R₂ is selected from the group consisting of hydrogen, alkyl of 1 toabout 4 carbon atoms, and alkenyl of about 2 to about 4 carbon atoms,

(d) R₃ is selected from the group consisting of ##STR5## where W isnitrogen or carbon;

(e) Het is selected from the group consisting of ##STR6## wherein (1) R₄is selected from the group consisting of

(a) R₁, --OR₁, --NHR₁, --S(O)_(n) R₁, and halogen, wherein n is 0, 1 or2, and R₁ is independently selected and as defined above, with theproviso that R₄ is not a camphor derivative or ##STR7## heterocyclogroup, (b) alkyl of 1 to about 12 carbon atoms substituted with Z₅wherein Z₅ is selected from the group consisting of hydroxy, halogen,--C(O)OH, --C(O)OR₈, --S(O)₃ OH, and --S(O)_(p) R₈ wherein R₈ is alkylof 1 to about 6 carbon atoms and p is 0, 1 or 2, and

(c) alkenyl of about 3 to about 6 carbon atoms;

(2) R₅ is selected from the group consisting of

(a) hydrogen,

(b) alkyl of 1 to about 10 carbon atoms,

(c) alkyl of 1 to about 3 carbon atoms substituted with cyclic alkyl ofabout 3 to about 8 carbon atoms,

(d) cyclic alkyl of 3 to about 6 carbon atoms,

(e) heterocycloalkyl of 4 to about 6 ring atoms with the ring atomsselected from carbon and heteroatoms, wherein the heteroatoms areselected from the group consisting of oxygen, nitrogen and --S(O)_(i) --wherein i is independently 0, 1 or 2,

(f) heterocyclo of 4 to about 6 ring atoms with the ring atoms selectedfrom carbon atoms and heteroatoms, wherein the heteroatoms are selectedfrom the group consisting of oxygen, nitrogen and --S(O)_(i) -- whereini is independently 0, 1 or 2 and which is attached to Het by a ringcarbon atom,

(g) alkenyl of 2 to about 6 carbon atoms which is optionally substitutedwith cyclic alkyl of 3 to about 5 carbon atoms,

(h) aryl which is optionally mono-, di- or tri-substituted with Y₁, Y₂and/or Y₃ respectively,

(i) heteroaryl of 5 to 6 atoms with the ring atoms selected from carbonatoms and heteroatoms, wherein the heteroatoms are selected from thegroup consisting of oxygen, nitrogen, and --S(O)_(i) -- wherein i isindependently 0, 1 or 2 and which is optionally mono-, di- ortri-substituted with Y₁, Y₂ and/or Y₃,

(j) aralkyl of about 7 to about 10 carbon atoms which is optionallymono-, di- or tri-substituted on the aryl ring with Y₁, Y₂ and/or Y₃ ;

(k) heteroaralkyl of 6 to 9 atoms with the ring atoms selected fromcarbon atoms and heteroatoms, wherein the heteroatoms are selected fromthe group consisting of oxygen, nitrogen and --S(O)_(i) -- wherein i isindependently 0, 1 or 2 and which is optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃,

(l) aralkenyl of 8 carbon atoms which is optionally mono-, di- ortri-substituted on the aryl ring with Y₁, Y₂ and/or Y₃,

(m) heteroaralkenyl of 7 to 8 atoms with the ring atoms selected fromcarbon atoms and heteroatoms, wherein the heteroatoms are selected fromthe group consisting of oxygen, nitrogen, and --S(O)_(i) -- wherein i isindependently 0, 1 or 2, and which is optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃,

(n) halogen,

(o) difluoromethyl or perfluoroalkyl of 1 to 3 carbon atoms,

(p) perfluorophenyl,

(q) perfluoroaralkyl of 7 to about 9 carbon atoms, and

(r) alkoxy of 1 to about 10 carbon atoms;

(3) R₆ is selected from the group consisting of

(a) R₁, --OR₁, --NHR₁, --S(O)_(n) R₁, and halogen, wherein n is 0, 1 or2, and R₁ is independently selected and as defined above, with theproviso that R₆ is not a camphor derivative or ##STR8## heterocyclgroup, and (b) alkyl of 1 to about 12 carbon atoms substituted with Z₆,wherein Z₆ is selected from the group consisting of hydroxy, halogen,--OR₉, --NHR₉, --C(O)OH, --C(O)OR₉, --S(O)₂ OH and --S(O)_(p) R₉ whereinR₉ is selected from alkyl of 1 to about 12 carbon atoms, aryl of about 6to about 10 carbon atoms optionally mono-, di- or tri-substituted on thering with Y₁, Y₂ and/or Y₃, aralkyl of about 7 to about 12 carbon atomsoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂ and/orY₃, heteroaryl of 1 to about 9 carbon atoms with the ring atoms selectedfrom carbon and heteroatoms selected from the group consisting ofoxygen, nitrogen and --S(O)_(p) -- and optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃ ; and heteroaralkyl ofabout 2 to about 10 carbon atoms with the ring atoms selected fromcarbon and heteroatoms selected from the group consisting of oxygen,nitrogen and --S(O)_(p) -- and optionally mono-, di- or tri-substitutedon the ring with Y₁, Y₂ and/or Y₃ ; and

(4) R₇ is independently selected from the R₅ group of substituents,provided that R₇ is not halogen; and pharmaceutically acceptable saltsthereof.

Peptidyl arginine aldehydes have been reported to exist in equilibriumstructures in aqueous solutions. Bajusz, S., et al., J. Med. Chem.,33:1729 (1990). These structures, as shown below, include the argininealdehyde, A, aldehyde hydrate, B, and two amino cyclol forms, C and D.The R group would represent the remainder of a given compound embodiedin the present invention. The peptide aldehydes of the present inventioninclude within their definition all the equilibrium forms. ##STR9##

Among other factors, the present invention is based on our finding thatthe novel compounds of our invention are active as selective inhibitorsof thrombin. In particular, we have found that certain of the preferredcompounds of the present invention exhibit advantageous selectivity inthat they are very potent inhibitors of thrombin but are inactive orsignificantly less active, (several orders of magnitude less) ininhibiting plasmin and are significantly less active in inhibitingtrypsin. This selectivity for inhibition of thrombin gives thesecompounds a therapeutic advantage in treating or preventing thrombosisin a mammal suspected of having a condition characterized by abnormalthrombosis.

In another aspect, the present invention is directed to pharmaceuticalcompositions comprising a therapeutically effective amount of a compoundof the present invention and a pharmaceutically acceptable carrier.

In yet another aspect, the present invention is directed to methods ofusing the compounds and pharmaceutical compositions of the presentinvention for the prevention of thrombosis in a mammal suspected ofhaving a condition characterized by abnormal thrombosis, comprisingadministering to said mammal a therapeutically effective amount of acompound of the present invention or pharmaceutical compositioncomprising such a compound.

DEFINITIONS

In accordance with the present invention and as used herein, thefollowing terms are defined to have following meanings, unlessexplicitly stated otherwise:

The term "alkenyl" refers to unsaturated aliphatic groups having atleast one double bond.

The term "alkyl" refers to saturated aliphatic groups includingstraight-chain, branched-chain and cyclic groups.

The terms "alkoxy" and "alkoxyl" refer to a group having the formula,R--O--, wherein R is an alkyl group.

The term "alkoxycarbonyl" refers to --C(O)OR wherein R is alkyl.

The term "aralkenyl" refers to an alkenyl group substituted with an arylgroup.

The term "aralkyl" refers to an alkyl group substituted with an arylgroup. Suitable aralkyl groups include benzyl, picolyl, and the like,all of which may be optionally substituted.

The term "aryl" refers to aromatic groups which have at least one ringhaving a conjugated pi electron system and includes carbocyclic aryl,heterocyclic aryl and biaryl groups, all of which may be optionallysubstituted.

The term "aryloxy" refers to a group having the formula, R--O--, whereinR is an aryl group.

The term "aralkoxy" refers to a group having the formula, R--O--,wherein R is an aralkyl group.

The term "amino acid" refers to natural amino acids, unnatural aminoacids, and amino acid analogs, all in their D and L stereoisomers iftheir structure allow such stereoisomeric forms. Natural amino acidsinclude alanine (Ala), arginine (Arg), asparagine (Asn), aspartic acid(Asp), cysteine (Cys), glutamine (Gln), glutamic acid (Glu), glycine(Gly), histidine (His), isoleucine (Ile), leucine (Leu), lysine (Lys),methionine (Met), phenylalanine (Phe), proline (Pro), serine (Ser),threonine (Thr), tryptophan (Trp), tyrosine (Tyr) and valine (Val).Unnatural amino acids include, but are not limited toazetidinecarboxylic acid, 2-aminoadipic acid, 3-aminoadipic acid,beta-alanine, aminopropionic acid, 2-aminobutyric acid, 4-aminobutyricacid, 6-aminocaproic acid, 2-aminoheptanoic acid, 2-aminoisobutyricacid, 3-aminoisobutyric acid, 2-aminopimelic acid, 2,4 diaminoisobutyricacid, desmosine, 2,2'-diaminopimelic acid, 2,3-diaminopropionic acid,N-ethylglycine, N-ethylasparagine, hydroxylysine, allo-hydroxylysine,3-hydroxyproline, 4-hydroxyproline, isodesmosine, alloisoleucine,N-methylglycine, N-methylisoleucine, N-methylvaline, norvaline,norleucine, ornithine and pipecolic acid. Amino acid analogs include thenatural and unnatural amino acids which are chemically blocked,reversibly or irreversibly, or modified on their N-terminal amino groupor their side-chain groups, as for example, methionine sulfoxide,methionine sulfone,

S-(carboxymethyl)-cysteine, S-(carboxymethyl)-cysteine sulfoxide andS-(carboxymethyl)-cysteine sulfone.

The term "amino acid analog" refers to an amino acid wherein either theC-terminal carboxy group, the N-terminal amino group or side-chainfunctional group has been chemically modified to another functionalgroup. For example, aspartic acid-(beta-methyl ester) is an amino acidanalog of aspartic acid; N-ethylglycine is an amino acid analog ofglycine; or alanine carboxamide is an amino acid analog of alanine.

The term "amino acid residue" refers to radicals having the structure:(1) --C(O)--R--NH--, wherein R typically is --CH(R')--, wherein R' is Hor a carbon containing substituent; or (2) ##STR10## wherein p is 1, 2or 3 representing the azetidinecarboxylic acid, proline or pipecolicacid residues, respectively.

"Biaryl" refers to phenyl substituted by carbocyclic or heterocyclicaryl as defined herein, ortho, meta or para to the point of attachmentof the phenyl ring.

"Brine" refers to an aqueous saturated solution of sodium chloride.

"Camphor derivative" refers to the groups: ##STR11##

"Carbocyclic aryl" refers to aromatic groups wherein the ring atoms ofthe aromatic ring are carbon atoms. Carbocyclic aryl groups includemonocyclic carbocyclic aryl groups, such as phenyl; naphthyl and otherpolycyclic groups, all of which may be optionally substituted. Suitablecarbocyclic aryl groups include phenyl and naphthyl. Suitablesubstituted carbocyclic aryl groups include indene and phenylsubstituted by one to two substituents such being advantageously loweralkyl, hydroxy, lower alkoxy, lower alkoxycarbonyl, halogen,trifluoromethyl, nitro, and cyano. Substituted naphthyl refers to 1- or2-naphthyl substituted by lower alkyl, lower alkoxy, or halogen.

"Cycloalkenyl" refers to a cyclic alkenyl group. Suitable cycloalkenylgroups include, for example, cyclopentenyl and cyclohexenyl.

"Cycloalkyl" refers to a cyclic alkyl group. Suitable cycloalkyl groupsinclude, for example, cyclohexyl, cyclopropyl, cyclopentyl, andcycloheptyl.

"Cyclohexylmethyl" refers to a cyclohexyl group attached to CH₂.

The term "halogen" refers to fluorine, chlorine, bromine and iodine.

"Heteroaralkenyl" refers to an alkenyl group substitued with aheteroaryl, and includes those heterocyclic systems described in"Handbook of Chemistry and Physics", 49th edition, 1968, R. C. Weast,editor;The Chemical Rubber Co., Cleveland, Ohio. See particularlySection C, Rules for Naming Organic Compounds, B. FundamentalHeterocyclic Systems.

"Heteroaralkyl" refers to an alkyl group substituted with a heteroaryl,and includes those heterocyclic systems described in "Handbook ofChemistry and Physics", 49th edition, 1968, R. C. Weast, editor;TheChemical Rubber Co., Cleveland, Ohio. See particularly Section C, Rulesfor Naming Organic Compounds, B. Fundamental Heterocyclic Systems.

"Heteroaryl" refers to aryl groups having from 1 to 9 carbon atoms andthe remainder of the atoms are heteroatoms, and includes thoseheterocyclic systems described in "Handbook of Chemistry and Physics",49th edition, 1968, R. C. Weast, editor;The Chemical Rubber Co.,Cleveland, Ohio. See particularly Section C, Rules for Naming OrganicCompounds, B. Fundamental Heterocyclic Systems. Suitable heteroatomsinclude oxygen, nitrogen, S(O)_(i), wherein i is 0, 1 or 2, and suitableheterocyclic aryls include furanyl, thienyl, pyridyl, pyrrolyl,pyrimidyl, pyrazinyl, imidazolyl, and the like.

"Heterocyclo" refers to a reduced heterocyclic ring system comprised ofcarbon, nitrogen, oxygen and/or sulfur atoms, and includes thoseheterocyclic systems described in "Handbook of Chemistry and Physics",49th edition, 1968, R. C. Weast, editor;The Chemical Rubber Co.,Cleveland, Ohio. See particularly Section C, Rules for Naming OrganicCompounds, B. Fundamental Heterocyclic Systems.

"Heterocycloalkyl" refers to an alkyl group substituted with aheterocyclo group, and includes those heterocyclic systems described in"Handbook of Chemistry and Physics", 49th edition, 1968, R. C. Weast,editor;The Chemical Rubber Co., Cleveland, Ohio. See particularlySection C, Rules for Naming Organic Compounds, B. FundamentalHeterocyclic Systems.

The term "lower" referred to herein in connection with organic radicalsor compounds defines such with up to and including 5, preferably up toand including 4 and advantageously one or two carbon atoms. Such groupsmay be straight chain or branched chain.

"Perfluoroalkyl" refers to an alkyl group which has every hydrogenreplaced with fluorine.

"Perfluoroaryl" refers to an aryl group which has every hydrogenreplaced with fluorine.

"Perfluoroaryl alkyl" refers an aralkyl group in which every hydrogen onthe aryl moiety is replaced with fluorine.

"Pharmaceutically acceptable salt" includes salts of the compounds ofthe present invention derived from the combination of such compounds andan organic or inorganic acid. In practice the use of the salt formamounts to use of the base form. The compounds of the present inventionare useful in both free base and salt form, with both forms beingconsidered as being within the scope of the present invention.

The term "Arg-al" refers to the residue of L-argininal which has theformula: ##STR12##

"N-alpha-t-butoxycarbonyl-N^(g) -nitro-L-arginine" refers to thecompound which has the formula: ##STR13##

In addition, the following abbreviations stand for the following:

"Boc" or "BOC" refers to t-butoxycarbonyl.

"BOP" refers to benzotriazol-1-yl-oxy-tris-(dimethylamino)-phosphoniumhexafluorophosphate.

"BzlSO₂ " refers to benzylsulfonyl.

"DCC" refers to N,N'-dicyclohexylcarbodiimide.

"EDC" refers to 1-ethyl-3-(3-dimethylaminopropyl)carbodiimidehydrochloride salt.

"HBTU" refers to 2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluroniumhexafluorophosphate.

"HCl" refers to hydrochloric acid.

"HOBt" refers to 1-hydroxybenzotriazole monohydrate.

"HPLC" refers to high pressure liquid chromatography.

"2-PrPen" refers to 2-propylpentanoyl.

"LiAlH₄ " refers to lithium aluminum hydride.

"LiAlH₂ (OEt)₂ refers to lithium aluminum dihydride diethoxide.

"NaOH" refers to sodium hydroxide.

"NMM" refers to N-methylmorpholine.

"TBTU" refers to 2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluroniumtetrafluoroborate.

"THF" refers to tetrahydrofuran.

"TLC" refers to thin layer chromatography.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts a general reaction scheme for preparation of certaincompounds of the present invention. In this figure, i) through iv) aredefined as: i) N-hydroxybenzotriazole, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride salt and N-methylmorpholine; ii) hydrogengas, palladium on carbon, ethanol, acetic acid and water; iii) 3 N HCl;and iv) sodium acetate, followed by HPLC purification using 0.1%trifluoroacetic acid in acetonitrile and water.

FIG. 2 depicts a general reaction scheme for preparation of certaincompounds of the present invention. In this figure, I) through viii) aredefined as: i) sodium hydride and ethyl bromoacetate; ii) hydrogen gasand palladium on carbon; iii) collidine and R₁ --SO₂ --Cl, where R₁ isas defined herein; iv) aqueous sodium hydroxide and methanol; v) N^(g)-nitro-L-argininal ethyl cyclol, hydrochloride salt,N-hydroxybenzotriazole, 1-ethyl-3-(3-dimethylamino-propyl)carbodiimidehydrochloride salt and N-methylmorpholine; vi) hydrogen gas, palladiumon carbon, ethanol, acetic acid and water; vii) 3N HCl; and viii) sodiumacetate, and then HPLC purification using 0.1% trifluoroacetic acid inacetonitrile and water.

FIG. 3 depicts a general reaction scheme for preparation of certaincompounds of the present invention. In this figure, I) through xiv) aredefined as: i) R₄ --C(═NH)--NH₂, where R₄ is as defined herein; ii)sodium hydride, allyl bromide; iii) sodium hydroxide; iv) triethylamine,diphenylphosphoryl azide and heat; v) t-butyl alcohol and heat; vi)trifluoroacetic acid; vii) collidine and R₁ --SO₂ --Cl, where R₁ is asdefined herein; viii) N-methylmorpholine-N-oxide and osmium tetroxide;ix) sodium periodate; x) sodium chlorite; xi) N^(g) -nitro-L-argininalethyl cyclol, hydrochloride salt, N-hydroxybenzotriazole,1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride salt andN-methylmorpholine; xii) palladium on carbon, ethanol, acetic acid andwater; xiii) 3N HCl; and xiv) sodium acetate, and then HPLC purificationusing 0.1% trifluoroacetic acid in acetonitrile and water.

FIG. 4 depicts a general reaction scheme for preparation of certaincompounds of the present invention. In this figure, i) though x) aredefined as: i) 1,1,1-3,3,3-hexamethyldisilazane andchlorotrimethylsilane; ii) R₇ X heated in dimethylformamide, wherein R₇is as defined herein and X is a halogen; iii) tetrabutylammoniumfluoride and ethyl bromoacetate; iv) hydrogen gas and palladium oncarbon; v) collidine and R₁ --SO₂ --Cl, where R₁ is as defined herein;vi) sodium hydroxide; vii) N^(g) -nitro-L-argininal ethyl cyclol,hydrochloride salt, N-hydroxybenzotriazole,1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride salt andN-methylmorpholine; viii) palladium on carbon, ethanol, acetic acid andwater; ix) 3N HCl; and x) sodium acetate, and then HPLC purificationusing 0.1% trifluoroacetic acid in acetonitrile and water.

FIG. 5 depicts the reaction scheme for preparation of a compound of thepresent invention. In this figure, "Boc" refers to the protecting group,t-butoxycarbonyl; "Cbz" refers to the protecting group,benzyloxycarbonyl; and "t-Bu" refers to the protecting group, t-butyl.Also, in this figure, i) through x) are defined as: i) lithium aluminumhydride; ii) ethanol and HCl, iii) hydrogen gas and palladium on carbon,1.0N HCl; iv) sodium hydride and t-butyl bromoacetate; v) hydrogen gasand palladium on carbon; vi) sodium bicarbonate and allyl chloroformate;vii) trifluoroacetic acid; viii) N-hydroxybenzotriazole,1-ethyl-3-(3-dimethylamino-propyl)carbodiimide hydrochloride salt andN-methylmorpholine; ix) hexafluorophosphoric acid; and x) sodium acetateand then HPLC purification using 0.1% trifluoroacetic acid inacetonitrile and water.

FIG. 6 depicts the anticoagulant effect of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl] acetyl-L-arginalmeasured in citrated human plasma, closed circles (1), using theactivated partial thromboplastin time (APTT) assay. The control clottingtimes (0 inhibitor) for human plasma was 29 seconds. The concentrationof this compound of the present invention which caused a doubling of thecontrol clotting time in human plasma was 7.7 micromolar. The data isthe mean of two independent determinations.

FIG. 7 depicts a reaction scheme for the preparation of compoundswherein X is a direct link. In this figure, i) through ix) are definedas i) hydrogen gas and palladium on carbon; ii) di-t-butyldicarbonateand sodium bicarbonate; iii) sodium hydride and R₁ iodide; iv) sodiumhydroxide; v) EDC, HOBt, and N-methylmorpholine; vi) hydrogen gas andpalladium on carbon, ethanol, acetic acid and water; vii) 3Nhydrochloric acid; and viii) sodium acetate, and then HPLC purificationusing 0.1% trifluoroacetic acid in acetonitrile and water.

FIG. 8 depicts a reaction scheme for the preparation of certaincompounds of the present invention. In this Figure, i) through vi) are:i) potassium carbonate, dimethylformamide and t-butyl bromoacetate; ii)lithium hydroxide and tetrahydrofuran; iii) triethylamine,diphenylphosphoryl azide, dioxane and heat (about ₋₋ ° C.); benzylalcohol and Δ; iv) hydrogen gas and palladium on carbon; v) R₁ SO₂ Cland collidine; and vi) trifluoroacetic acid. See also Examples 91-96. R₁is as defined in connnection with formula (I) herein.

FIG. 9 depicts a reaction scheme for the preparation of certaincompounds of the present invention, including compound 18 in FIG. 3. Inthis Figure, i) through vi) are: i) tetra-n-butyl ammonium fluoride,dimethoxyethane, and t-butyl bromoacetate; ii) lithium hydroxide andtetrahydrofuran; iii) triethylamine, diphenylphosphorylazide, dioxaneand Δ; benzyl alcohol and Δ; iv) hydrogen gas and palladium on carbon;v) R₁ SO₂ Cl and 4-methylmorpholine and vi) trifluoroacetic acid. Seealso Examples 102-107. R₁ and R₄ are as defined in connection withformula (I) herein.

FIG. 10 depicts a reaction scheme for the preparation of certaincompounds of the present invention, including compound 26 in FIG. 4. Inthis Figure, i) through v) are: i) potassium carbonate, R₇ X anddimethylsulfoxide; ii) sodium hydride and t-butylbromoacetate; iii)hydrogen gas and 10% palladium on carbon; iv) R₁ SO₂ Cl and4-methylmorpholine; and v) trifluoroacetic acid. See also Examples108-111. R₁ and R₇ are as defined in connection with formula (I) herein.

FIG. 11 depicts a reaction scheme for the preparation of certaincompounds of the present invention. In this Figure, i) through vii) are:i) 2 equivalents, lithium diisoprophylamide, R_(x) X, 50% sulfuric acid;iii) triethylamine, diphenylphosphonyl azide, dioxane and Δ; benzylalcohol and Δ; iv) sodium hydride, dimethylformamide and t-butylbromoacetate; v) hydrogen gas and palladium on carbon; vi) R₁ SO₂ Cl andcollidine; and vii) trifluoroacetic acid. See also Examples 97-101. R₁is as defined in connection with formula (I) herein. R_(x) is any R₄substituent minus one carbon, such as methyl if R₁ was ethyl and X ishalogen.

FIG. 12 depicts a reaction scheme for the preparation of certaincompounds of the present invention. In this Figure, i) through iv) are:i) lithium hexamethyldisilazide, chlorotrimethylsilane, lithiumhexamethyldisilazide and benzaldehyde; ii) lithium hexamethyldisilazideand ethyl bromoacetate; iii) acetic anhydride, 10% palladium on carbon,hydrogen gas; and iv) lithium hydroxide. See also Examples 114-116.

FIG. 13 depicts a reaction scheme for the preparation of certaincompounds of the present invention. In this Figure, i) through ix) are:i) thiourea and ethanol to give a 97% yield of 72 (compound of Example135); ii) chlorine gas, water to give 90% yield of 73 (compound ofExample 136); iii) lithium bis(trimethylsilyl)amide, tetrahydrofuran andt-butyl bromoacetate to give 75% yield of 75 (compound of Example 137);iv) hydrogen gas and palladium on carbon to give 98.5% yield of 76(compound of Example 138); v) 4-methylmorpholine, and acetonitrile togive 65% yield of 78 (compound of Example 139); vi) trifluoroacetic acidand methylene chloride to give a quantitative yield of 79 (compound ofExample 140). vii) N-hydroxybenzotriazole,1-ethyl-3-(3-dimethylamino-propyl)carbodiimide hydrochloride salt, 80(N^(g) -nitro-L-argininal ethyl cyclol), 4-methylmorpholine at roomtemperature to give 55% yield of 81 (compound of Example 141). viii)hydrogen gas (1 atm), palladium on carbon, ethanol, acetic acid to givequantitative yield of 82 (compound of Example 142); and ix) 6N HCl, roomtermperature, about one hour to give 74% yield of 83 (compound ofExample 143). See also Examples 135 to 143.

DETAILED DESCRIPTION OF THE INVENTION

1. Preferred Compounds

Compounds of the present invention have the formula: ##STR14## wherein

(a) X is selected from the group consisting of --S(O)₂ --,--N(R')--S(O)₂ --, --(C═O)--, --OC(═O)--, --NH--C(═O)--, --P(O) (R")--and a direct link, wherein R' is hydrogen, alkyl of 1 to about 4 carbonatoms, aryl of about 6 to about 14 carbon atoms or aralkyl of about 6 toabout 16 carbon atoms, and R" is NR', OR', R', or SR', with the provisothat R" is not NH, OH, H, or SH, and;

(b) R₁ is selected from the group consisting of:

(1) alkyl of 1 to about 12 carbon atoms,

(2) alkyl of 1 to about 3 carbon atoms substituted with cyclic alkyl ofabout 3 to about 8 carbon atoms, which optionally is substituted in thering carbons with hydroxyl, amino, guanidino, amidino, or alkoxyl oralkyl each of 1 to about 3 carbons,

(3) cyclic alkyl of 3 to about 15 carbon atoms, which optionally issubstituted in the ring carbons with hydroxyl, amino, guanidino,amidino, or alkoxyl or alkyl each of 1 to about 3 carbons,

(4) heterocycloalkyl of 4 to about 10 ring atoms with the ring atomsselected from carbon and heteroatoms, wherein the heteroatoms areselected from the group consisting of oxygen, nitrogen, and S(O)_(i),wherein i is 0, 1 or 2, and which is optionally substituted on the ringcarbons with hydroxyl, alkoxyl or alkyl each of 1 to about 3 carbons,amino, guanidino, or amidino,

(5) heterocyclo of 4 to about 10 ring atoms with the ring atoms selectedfrom carbon and heteroatoms, wherein the heteroatoms are selected fromthe group consisting of oxygen, nitrogen, and S(O)_(i), wherein i is 0,1 or 2, including the group ##STR15## wherein ##STR16## is a 5 to 7member heterocycle of 3 to 6 ring carbon atoms, where V is --CH₂ --,--O--, --S(═O)--, --S(O)₂ -- or --S--, and which is optionallysubstituted on the ring carbons with hydroxyl, alkoxyl, or alkyl each of1 to about 3 carbons, amino, guanidino, or amidino,

(6) alkenyl of 2 to about 6 carbon atoms which is optionally substitutedwith cyclic alkyl of about 3 to about 8 carbon atoms, which optionallyis substituted in the ring carbons with hydroxyl, amino, guanidino,amidino, or alkoxyl or alkyl each of 1 to about 3 carbons,

(7) aryl of about 6 to about 14 carbon atoms which is optionally mono-,di- or tri-substituted with Y₁, Y₂, and/or Y₃,

(8) heteroaryl of 5 to 14 atoms with the ring atoms selected from carbonand heteroatoms, wherein the heteroatoms are selected from oxygen,nitrogen, and S(O)_(i), wherein i is 0, 1 or 2, and which is optionallymono-, di- or tri-substituted with Y₁, Y₂, and/or Y₃,

(9) aralkyl of about 7 to about 15 carbon atoms which is optionallysubstituted on the alkyl chain with hydroxy or halogen and optionallymono-, di-, or tri-substituted on the aryl ring with Y₁, Y₂, and/or Y₃,

(10) heteroaralkyl of 6 to 11 atoms with the ring atoms selected fromcarbon and heteroatoms, wherein the heteroatoms are selected fromoxygen, nitrogen, and S(O)_(i), wherein i is 0, 1 or 2, and which isoptionally substituted on the alkyl chain with hydroxy or halogen andoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂, and/orY₃,

(11) aralkenyl of about 8 to about 16 carbon atoms which is optionallymono-, di-, or tri-substituted on the aryl ring with Y₁, Y₂, and/or Y₃,

(12) heteroaralkenyl of 7 to 12 atoms with the ring atoms selected fromcarbon and heteroatoms, wherein the heteroatoms are selected fromoxygen, nitrogen, and S(o)_(i), wherein i is 0, 1 or 2, and which isoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂, and/orY₃, respectively, ##STR17## (17) difluoromethyl and perfluoroalkyl of 1to about 12 carbon atoms, (18) perfluoroaryl of about 6 to about 14carbon atoms,

(19) perfluoroaralkyl of about 7 to about 15 carbon atoms, and

(20) hydrogen,

wherein Y₁, Y₂, and Y₃ are

(i) independently selected from the group consisting of hydrogen,halogen, cyano, tetrazolyl, amino, guanidino, amidino, methylamino, andmethylguanidino, --CF₃, --CF₂ H, --CF₂ CF₃, --CH(CF₃)₂, --C(OH)(CF₃)₂,--OCF₃, --OCF₂ CF₃, --OC(O)NH₂, --OC(O)NHZ₁, --OC(O)NZ₁ Z₂, --NHC(O)Z₁,--NHC(O)NH₂, --NHC(O)NZ₁, --NHC(O)NZ₁ Z₂, --C(O)OH, --C(O)NH₂,--C(O)NHZ₁, --C(O)OZ₁, --P(O)₃ H₂, --P(O)₃ (Z₁)₂, --S(O)₃ H, --S(O)_(m)Z₁, --Z₁, --OZ₁, --OH, --NH₂, --NHZ₁, and --NZ₁ Z₂, wherein m is 0, 1 or2, and Z₁ and Z₂ are independently selected from the group consisting ofalkyl of 1 to about 12 carbon atoms, aryl of about 6 to about 14 carbonatoms, heteroaryl of about 5 to about 14 atoms having 1 to about 9carbon atoms, aralkyl of about 7 to about 15 carbon atoms, andheteroaralkyl of about 6 to about 11 atoms having about 3 to about 9carbon atoms, or

(ii) Y₁ and Y₂ are selected together to be --OC(Z₃) (Z₄) O--, wherein Z₃and Z₄ are independently selected from the group consisting of hydrogen,alkyl of 1 to about 12 carbon atoms, aryl of about 6 to about 14 carbonatoms, heteroaryl of about 5 to about 14 atoms having 1 to about 9carbon atoms, aralkyl of about 7 to about 15 carbon atoms, andheteroaralkyl of about 6 to about 11 atoms having about 3 to about 9carbon atoms, with the proviso that if X is not a direct link, then R₁is not hydrogen;

(c) R₂ is selected from the group consisting of hydrogen, alkyl of 1 toabout 4 carbon atoms, and alkenyl of 2 to about 4 carbon atoms,

(d) R₃ is selected from the group consisting of ##STR18## where W isnitrogen or carbon;

(e) Het is selected from the group consisting of ##STR19## wherein (1)R₄ is selected from the group consisting of

(a) R₁, --OR₁, --NHR₁, --S(O)_(n) R₁, and halogen, wherein n is 0, 1 or2, and R₁ is independently selected and as defined above, with theproviso that R₄ is not a camphor derivative or ##STR20## heterocyclgroup, (b) alkyl of 1 to about 12 carbon atoms substituted with Z₅wherein Z₅ is selected from the group consisting of hydroxy, halogen,--C(O)OH, --C(O)OR₈, --S(O)₃ OH, and --S(O)_(p) R₈ wherein R₈ is alkylof 1 to about 6 carbon atoms and p is 0, 1 or 2, and

(c) alkenyl of about 3 to about 6 carbon atoms;

(2) R₅ is selected from the group consisting of

(a) hydrogen,

(b) alkyl of 1 to about 10 carbon atoms,

(c) alkyl of 1 to about 3 carbon atoms substituted with cyclic alkyl ofabout 3 to about 8 carbon atoms,

(d) cyclic alkyl of 3 to about 6 carbon atoms,

(e) heterocycloalkyl of 4 to about 6 ring atoms with the ring atomsselected from carbon and heteroatoms, wherein the heteroatoms areselected from the group consisting of oxygen, nitrogen and --S(O)_(i) --wherein i is independently 0, 1 or 2,

(f) heterocyclo of 4 to about 6 ring atoms with the ring atoms selectedfrom carbon atoms and heteroatoms, wherein the heteroatoms are selectedfrom the group consisting of oxygen, nitrogen and --S(O)_(i) -- whereini is independently 0, 1 or 2 and which is attached to Het by a ringcarbon atom,

(g) alkenyl of 2 to about 6 carbon atoms which is optionally substitutedwith cyclic alkyl of 3 to about 5 carbon atoms,

(h) aryl which is optionally mono-, di- or tri-substituted with Y₁, Y₂and/or Y₃ respectively,

(i) heteroaryl of 5 to 6 atoms with the ring atoms selected from carbonatoms and heteroatoms, wherein the heteroatoms are selected from thegroup consisting of oxygen, nitrogen, and --S(O)_(i) -- wherein i isindependently 0, 1 or 2 and which is optionally mono-, di- ortri-substituted with Y₁, Y₂ and/or Y₃,

(j) aralkyl of about 7 to about 10 carbon atoms which is optionallymono-, di- or tri-substituted on the aryl ring with Y₁, Y₂ and/or Y₃ ;

(k) heteroaralkyl of 6 to 9 atoms with the ring atoms selected fromcarbon atoms and heteroatoms, wherein the heteroatoms are selected fromthe group consisting of oxygen, nitrogen and --S(O)_(i) -- wherein i isindependently 0, 1 or 2 and which is optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃,

(l) aralkenyl of 8 carbon atoms which is optionally mono-, di- ortri-substituted on the aryl ring with Y₁, Y₂ and/or Y₃,

(m) heteroaralkenyl of 7 to 8 atoms with the ring atoms selected fromcarbon atoms and heteroatoms, wherein the heteroatoms are selected fromthe group consisting of oxygen, nitrogen, and --S(O)_(i) -- wherein i isindependently 0, 1 or 2, and which is optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃,

(n) halogen,

(o) difluoromethyl or perfluoroalkyl of 1 to 3 carbon atoms,

(p) perfluorophenyl,

(q) perfluoroaralkyl of 7 to about 9 carbon atoms, and

(r) alkoxy of 1 to about 10 carbon atoms;

(3) R₆ is selected from the group consisting of

(a) R₁, --OR₁, --NHR₁, --S(O)_(n) R₁, and halogen, wherein n is 0, 1 or2, and R₁ is independently selected and as defined above, with theproviso that R₆ is not a camphor derivative or ##STR21## heterocyclogroup, and (b) alkyl of 1 to about 12 carbon atoms substituted with Z₆,wherein Z₆ is selected from the group consisting of hydroxy, halogen,--OR₉, --NHR₉, --C(O)OH, --C(O)OR₉, --S(O)₂ OH and --S(O)_(p) R₉ whereinR₉ is selected from alkyl of 1 to about 12 carbon atoms, aryl of about 6to about 10 carbon atoms optionally mono-, di- or tri-substituted on thering with Y₁, Y₂ and/or Y₃, aralkyl of about 7 to about 12 carbon atomsoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂ and/orY₃, heteroaryl of 1 to about 9 carbon atoms with the ring atoms selectedfrom carbon and heteroatoms selected from the group consisting ofoxygen, nitrogen and --S(O)_(p) -- and optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃ ; and heteroaralkyl ofabout 2 to about 10 carbon atoms with the ring atoms selected fromcarbon and heteroatoms selected from the group consisting of oxygen,nitrogen and --S(O)_(p) -- and optionally mono-, di- or tri-substitutedon the ring with Y₁, Y₂ and/or Y₃ ; and

(4) R₇ is independently selected from the R₅ group of substituents,provided that R₇ is not halogen; and pharmaceutically acceptable saltsthereof.

Preferred X groups include --SO₂ --, --NH--S(O)₂ --, and --N(R')--S(O)₂--. Especially preferred X groups include --SO₂ --.

Preferred R₁ groups include alkyl, aralkyl, and aryl groups. PreferredR₁ aryl groups include substituted or unsubstituted phenyl and naphthyl.Preferred substitutions include, methyl, methoxy, fluoro, chloro,trifluoromethyl, and --OCF₃. Meta and ortho substitution is preferred.

Particularly preferred R₁ groups include aralkyl groups. Especiallypreferred R₁ groups include substituted or unsubstituted benzyl andnaphthyl groups. Cyclohexyl and cyclohexylmethyl are other especiallypreferred R₁ groups.

A particularly preferred R₂ group is hydrogen.

Preferred R₃ groups include ##STR22##

Preferred R₄ groups include:

(i) hydrogen,

(ii) alkyl of 1 to 6 carbon atoms or alkyl of 1 to 6 carbon atomssubstituted with Z₅, wherein Z₅ is selected from the group consisting ofhydroxy, halogen, --C(O)OH, --C(O)OR₈, --S(O)₃ OH and --S(O)_(p) R₈wherein R₈ is alkyl of 1 to about 6 carbon atoms, and p is 0, 1 or 2,

(iii) alkyl of 1 to 3 carbon atoms substituted with cyclic alkyl of 3 to5 carbon atoms,

(iv) alkenyl of about 3 to about 6 carbon atoms,

(v) cycloalkyl of about 3 to about 5 carbon atoms,

(vi) heteroaryl of 5 atoms, and

(vii) heteroaralkyl of 6 atoms.

Preferred R₅ groups include hydrogen, halogen, alkyl of 1 to about 5carbon atoms, trifluoromethyl, and alkoxy of 1 to 4 carbon atoms.Hydrogen is an especially preferred R₅ group.

Preferred R₆ groups include:

(i) hydrogen,

(ii) alkyl of 1 to about 12 carbon atoms or alkyl of 1 to 12 carbonatoms substituted with Z₆, wherein Z₆ is selected from the groupconsisting of hydroxy, halogen, --OR₉, --NHR₉ --C(O)OH, --C(O)OR₉,--S(O)₂ OH and --S(O)_(p) R₉, wherein R₉ is as defined above;

(iii) alkyl of 1 to about 3 carbon atoms substituted with cyclic alkylof about 6 to about 8 carbon atoms;

(iv) alkenyl of 2 to about 6 carbon atoms which is optionallysubstituted with cyclic alkyl of about 3 to about 8 carbon atoms or arylof about 3 to about 10 carbon atoms;

(v) aralkyl or substituted aralkyl, as defined above;

(vi) heteroaralkyl or substituted aralkyl, as defined above;

(vii) aralkenyl of about 8 to 15 carbon atoms which is optionally mono-,di- or tri-substituted on the ring with Y₁, Y₂ and/or Y₃, as definedabove;

(viii) heteroaralkenyl or substituted heteroaralkenyl, as defined above.

More preferred R₆ groups, when R₄ and R₅ are hydrogen or methyl, areselected from the group consisting of aralkyl of about 8 to about 13carbon atoms, and --O-- aralkyl, --NH--aralkyl, and --S(O)_(p) -aralkylof about 7 to about 12 carbon atoms. Preferred aryl portions of thearalkyl groups include unsubstituted and substituted phenyl or naphthyl.Preferred substitutions on the aryl ring include methyl, methoxy,fluoro, chloro and trifluoromethyl. Phenylethyl, phenylpropyl, hydrogen,cyclohexylethyl and cyclohexylpropyl are especially preferred R₆ groups.

Preferred R₇ groups include hydrogen, methyl, difluoromethyl andtrifluoromethyl. Hydrogen is an especially preferred R₇ group.

Preferred Het groups include ##STR23##

A particularly preferred Het, when R₅ and R₆ are independently selectedto be hydrogen or methyl, is ##STR24## wherein R₄ is selected from thegroup consisting of hydrogen, methyl, ethyl, propenyl, allyl, propyl,isopropyl, butyl, R-sec-butyl, S-sec-butyl, isobutyl, 1-pentyl,R-2-pentyl, S-2-pentyl, 3-pentyl, S-1-(2-methyl)-butyl,R-2-(3-methyl)-butyl, 1-(3-methyl)-butyl, R-1-(2-methyl)-butyl,cyclopentyl, 2-pyrolyl, 3-pyrolyl, 1-hexyl, S-2-hexyl, R-2-hexyl,R-3-hexyl, and S-3-hexyl. A particularly preferred Het according to thisaspect has hydrogen or methyl as R₄.

According to a particularly preferred aspect, provided are compounds offormula I wherein X is --S(O)₂ --, R₁ is substituted or unsubstitutedaryl or aralkyl, R₃ is ##STR25## and Het is ##STR26## A very preferredaspect is directed to such compounds where R₁ is substituted orunsubstituted benzyl or phenyl.

Preferred compounds include

3-[(phenylsulfonyl)amino-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 90, Compound B),

3-[(2-naphthylsulfonyl)amino]-2-oxo-1,2 dihydropyridylacetyl-L-argininal(Example 90),

3-[(1-naphthylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 90),

3-(cyclohexylaminosulfonylamino-2-oxo-1,2-dihydropyridyl)-acetyl-L-argininal(Example 90),

3-(phenylaminosulfonylamino-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(phenoxycarbonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(cyclohexylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(cyclohexylmethylsulfonyl)amino]-2-oxo-1,2dihydropyridylacetyl-L-argininal (Example 121H),

3-[(phenethylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 121G),

3-[(2-methoxycarbonylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(3-methoxycarbonylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(4-methoxycarbonylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(2-trifluoromethylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 90),

3-[(3-trifluoromethylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 90),

3-[(4-trifluoromethylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 90),

3-[(2-methoxycarbonylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(3-methoxycarbonylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(4-methoxycarbonylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

3-[(2-trifluoromethylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 90),

3-[(3-trifluoromethylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal(Example 121L),

3-[(4-trifluoromethylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal(Example 10),

[3-[(benzylsulfonyl)amino]-6-methyl-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal(Example 19 and Example 113, Compound C),

5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinyl-acetyl-L-argininal(Example 29b),

2-methyl-5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetyl-L-argininal(Example 40 and 113, Compound D),

5-benzylsulfonylamino-uracilylacetyl-L-argininal,

5-benzylsulfonylamino-1-methyl-uracilylacetyl-L-argininal (Example 54and Example 113, Compound E),

3-[(2-trifluoromethylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,

[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-3-[3-piperidyl-(N-guanidino)]alaninal,and

[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-D,L-3-amidinophenylalaninal.

According to another aspect, the present invention is directed to saltsof the compounds of formula (I). "Salt" includes within its definition,salts of the compounds of the present invention derived from thecombination of such compounds and an organic or inorganic acid. Inpractice, the use of the salt form amounts to use of the base form. Thecompounds of the present invention are useful in both free base and saltform, with both forms being considered as being within the scope of thepresent invention. These salts include acid addition salts, for example,salts of hydrochloric acid, hydrobromic acid, acetic acid, benzenesulfonic acid and other suitable acid addition salts.

2. Preparation of Preferred Compounds

FIG. 1 exemplifies a preferred reaction scheme for the synthesis ofcertain compounds of the present invention. N^(g) -nitro-L-argininalethyl cyclol, hydrochloride salt 2 is coupled to the terminal carboxylof the R₁ sulfonyl amino heterocycle 1 to give 3. Especially preferredcoupling reagents are N-hydroxybenzotriazole in acetonitrile with1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride salt. 3 ishydrogenated with hydrogen gas and palladium on carbon to remove theN^(g) -nitro group to give 4. 4 is then treated with strong acid andpurified by HPLC with trifluoroacetic acid in the solvent to produceargininal 5.

The compounds of the present invention may be prepared by the preferredreaction schemes depicted in FIGS. 2 through 5. Examples 5 through 10provide the details of the preferred reaction scheme of FIG. 2, Examples20 through 29 provide the details for the preferred reaction scheme ofFIG. 3, Examples 41 through 47 provide the details for the preferredreaction scheme of FIG. 4, and Examples 55 through 63 provide thedetails for the preferred scheme of FIG. 5.

In these reaction schemes, intermediates, which include 9, 18, 26, and36 shown in FIGS. 2 through 5, respectively, are coupled to argininal orargininal mimic moieties to eventually give the compounds of the presentinvention. Examples 1 through 4 provide the details for the preparationof the precursor to the argininal moiety used in FIGS. 2 through 4.Examples 55 through 57 provide the details for the preparation of theargininal precursor used when hydrogenation sensitive groups exist.Examples 64 through 71 provide the details for the preparation ofcompounds of the present invention possessing a3-[3-piperidyl-(N-guanidino)]alaninal in the P₁ position.

The preferred means of chemically coupling (as for example, 9 to 10 ofFIG. 2 or 18 to 19 of FIG. 3) include formation of a peptide bond byusing conventional coupling reagents known in the art. See Bodanszky,N., Peptide Chemistry, pp. 55-73, Springer-Verlag, New York (1988) andreferences cited therein. The chemical coupling may be either by meansof one-step or two-step coupling. In one-step coupling, the two couplingpartners are coupled directly. Preferred coupling reagents for one-stepcoupling of the include DCC with HOBt, EDC with HOBt, HBTU or TBTU. Intwo-step coupling, an activated ester or anhydride of the C-terminalcarboxy group of one coupling partner is formed prior to its coupling tothe other coupling partner.

For example, as shown in FIG. 2, the nitrogen of the pyridine ring of 6is alkylated to give 7. The nitro group is then reduced to the amine,which is then reacted with a sulfonyl chloride, depicted by R₁ --S(O)₂--Cl, to give 8. R₁ is as defined herein. The ethyl ester of 8 isremoved by treatment with aqueous sodium hydroxide in methanol to givethe carboxylic acid 9. The acid of 9 is coupled to N^(g)-nitro-L-argininal ethyl cyclol HCl salt by carbodiimide coupling togive 10. 6-Alkylated pyridyl compounds are made according to Examples 11through 19. 10 is hydrogenated with hydrogen gas and palladium on carbonto remove the N^(g) -nitro group to give 11. 11 is hydrolyzed in aqueousacid to give 12.

FIG. 3 provides a preferred reaction scheme for preparing pyrimidylcompounds of the present invention. Examples 21 through 29 describe thispreparation. Pyrimidine 14 is alkylated with allyl bromide, and then theester is hydrolyzed with sodium hydroxide in methanol to give the1-allyl pyrimidone 15. 15 is then treated with triethylamine anddiphenylphosphoryl azide to form the acyl azide which undergoes theCurtius rearrangement. Reaction with t-butanol forms the BOC protected5-aminopyrimidone 16. Treatment with acid removes the BOC group. Theamine is then reacted with an alkyl sulfonyl chloride to give 17. 17 isoxidized in three steps to form 18, which undergoes coupling aspreviously described.

FIG. 9 provides an alternate preferred reaction scheme for preparingintermediate compound 18 of FIG. 3. Synthesis of 18 by this alternateroute is as described in Examples 102 to 107.

FIG. 4 provides a preferred reaction scheme for preparing uracilcompounds of the present invention. Examples 41 through 54 describe thispreparation. As shown in FIG. 4, 5-nitrouracil 22 is reacted with1,1,1,3,3,3-hexamethyldisilazane and chlorotrimethylsilane to give the5-nitrouracil bis(trimethylsilyl) ether, which is then reacted withbromomethylmethyl ether to give the methoxymethyl uracil 23. Thiscompound is then reacted with ethyl bromoacetate to give the ethyluracilylacetate 24. The nitro group is then reduced to the amine usinghydrogen gas and palladium on carbon. The amine is then treated with2,4,6-collidine and R₁ SO₂ Cl to give the amide 25. The ethyl ester isconverted to acid 26 by treatment with sodium hydroxide in methanol. Theacid of 26 is coupled to N^(g) -nitro-L-argininal ethyl cyclolhydrochloride salt (prepared according to Examples 1 through 4). Theadduct 27 is deprotected by treatment with hydrogen gas and palladium oncarbon in an ethanol, acetic acid, and water mixture. 28 is hydrolizedwith 3N hydrochloric acid and then purified by HPLC with a solventcontaining 0.1% trifluoroacetic acid to give argininal 29.

FIG. 10 provides an alternate preferred reaction scheme for preparingintermediate compound 26 of FIG. 4. Synthesis of 26 by this alternateroute is as described in Examples 108 to 111.

FIG. 5 provides a preferred reaction scheme for preparing compounds ofthe invention possessing a hydrogenation sensitive moiety in the P₄position. This method uses the di-N-t-butoxycarbonyl protecting groupfor the L-argininal moiety. This scheme has an alkenyl carbamate as thehydrogenation sensitive moiety. Examples 55 through 63 describe thispreparation which uses hexafluorophosphoric acid to remove the BOCprotecting groups. This general method can be used to prepare otherhydrogenation sensitive compounds.

As described by Example 58, 3-nitro-2-hydroxypyridine 33 is treated withsodium hydride and then t-butyl bromoacetate to give 34. The nitro groupof 34 is reduced to the amine by treatment with hydrogen gas andpalladium on carbon. The amine is condensed with allyl chloroformate inthe presence of sodium bicarbonate to give 35. The t-butyl group of 35is removed by trifluoroacetic acid to give 36.Alpha-N-t-benzyloxycarbonyl-omega, omega'-di-N-t-butoxycarbonylarginineis dissolved in acetonitrile and treated with hydroxybenzotriazole and1-ethyl-3-(3-dimethylaminopropyl)carbodiimide HCl salt to formalpha-N-benzyloxycarbonyl-omega, omega'-di-N-t-butoxycarbonyl-L-argininelactam. The lactam 30 is opened by treatment with LiAlH₄ intetrahydrofuran at -70° C. to provide alpha-N-benzyloxycarbonyl-omega,omega'-di-N-t-butoxycarbonyl-L-argininal 31. This aldehyde is protectedas the diethyl acetal by treatment with ethanol and HCl. TheN-benzyloxycarbonyl protecting group is removed by treatment withhydrogen gas and palladium on carbon to give omega,omega'-di-N-t-butoxycarbonyl-L-argininal diethyl acetal, HCl salt 32.This protected L-argininal moiety can then be coupled to a desiredcarboxylic acid, shown in the figure as 36, by treatment withN-hydroxybenzotriazole and1-ethyl-3-(3-dimethylamino-propyl)carbodiimide HCl salt. The diethylacetal and the di-BOC protecting groups are removed by treatment withhexafluorophosphoric acid in acetonitrile at 0° C. The reaction isquenched by adjusting to pH 4 with 2.5 M aqueous sodium acetate.Preparative HPLC using 0.1% CF₃ COOH in 10-40% aqueous acetonitrileprovides the trifluoroacetate salt of the desired substitutedL-argininal compound 38.

For preparation of certain compounds having hydrogenation sensitivesubstituent groups, it is preferred to avoid the use of hydrogen gaswith palladium on carbon. Another preferred method for preparingcompounds of the present invention containing hydrogenation sensitivegroups such as alkenyl or aryl moieties substituted with halogen, cyano,nitro, or --S--Z₁, is to use boron tris(trifluoroacetate), B(OCOCF₃)₃,to cleave the N^(g) -nitro of the arginine group. The reagent isprepared by the reaction of BBr₃ and CF₃ COOH in dichloromethane at 0°C. The reagent is also commercially available. Generally, the N^(g)-nitro compound is treated with boron tris(trifluoroacetate) intrifluoroacetic acid at 0° C. See, e.g., Fieser, M. and Fieser, L. F.,Reagents for Organic Synthesis, p. 46, John Wiley & Sons, New York(1974); Pless, J., and Bauer, W. Angew. Chem., Internat. Ed., 12, 147(1973).

In addition, another preferred reagent for selective nitro groupcleavage is titanium trichloride. This reagent is commerciallyavailable. The Ng nitro compound is treated with titanium trichloride inaqueous methanol containing an ammonium acetate buffer followed byexposure of the reaction mixture to air or dimethyl sulfoxide.Freidinger, R. M., Hirschmann, R., and Veber, D. F., J. Org. Chem., 43,4800 (1978).

FIG. 7 illustrates a preferred reaction scheme for the preparation ofcompounds where X is a direct link. This figure is described by Examples83 through 88.

As shown in FIG. 7, the nitro group of pyridone 7 is reduced bytreatment with hydrogen gas and palladium on carbon. The amine is thenprotected by the Boc group to form 39. The Boc protected amine 39 isthen treated with sodium hydride and alkylated with R₁ iodide, where R₁is as defined herein. The ethyl ester is converted to acid 40 by sodiumhydroxide. Acid 40 is then coupled to the compound of Example 4 bystandard coupling techniques using1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride salt,1-hydroxybenzotriazole monohydrate, and N-methylmorpholine to give 41.The N^(g) -nitro group is removed by catalytic hydrogenation withpalladium on carbon to give 42. The Boc protecting group is removed andthe argininal is unmasked by treatment with HCl, followed by sodiumacetate. HPLC purification with 0.1% trifluoroacetic acid gives thefinal product 43 in FIG. 7.

FIG. 8 provides an alternate reaction scheme for preparing theintermediate compound described in Example 16. Examples 91-96 describethis alternate synthetic route.

FIG. 11 provides a preferred reaction scheme for the preparation of6-substituted pyridones. Examples 97-101 describe this synthetic route.

FIG. 12 provides a preferred reaction scheme for the preparation of4-(hydroxyl substituted)alkyl or aralkyl pyridones. Examples 113-115describe this synthetic route.

FIG. 13 provides a reaction scheme for a preferred compound of thepresent invention,[3-(2-fluorobenzylsulfonyl)amino-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,Examples 135 to 143 describe this synthetic route. An overall yield of19% compound 83 was obtained.

Another aspect of the present invention is a method for alkylating a3-nitro-2-oxo-1,2-dihydropyridyl acetate compound at ring position 4comprising

(a) combining the compound with a solution of a zinc salt and an alkylgrignard under anhydrous conditions to form a3-nitro-2-oxo-4-alkyl-1,2,3,4-dihydropyridyl acetate intermediate,

(b) contacting the intermediate with an oxidizing agent, and

(c) recovering a 4-alkyl-3-nitro-2-xo-1,2-dihydropyridyl acetateproduct.

The disclosed method provides an efficient route to the synthesis of anintermediate used int he synthesis of compounds having a 6-memberedheterocycle comprising an alkyl at the four position and a nitro at thethree position. Example 134 exemplifies several such compounds.

Zinc salts such as zinc chloride, zinc bromide, and zinc iodide can beused in step (a). Zinc chloride and zinc bromide are preferred, and zincchloride is especially preferred.

An alkyl grignard results from reacting a magnesium metal with an alkylor aryl bromide, chloride, or idodide. The magnesium inserts betwen thehalogen and the carbon bond to form the grignard. According to thisaspect of the invention, the alkyl grignard in (a) is synthesized from astarting compound in the group defined by [R1]. A preferred alkylgrignard is 3-phenylpropyl magnesium bromide.

Step (a) can be performed by combining the compund and the zinc salt,follwed by addition of the alkyl grignard, or by combining the zinc saltand the grignard, followed by adition of the compound. Preferred is amethod wherien the compound and the zinc salt are combined, followed byaddition of the alkyl grignard.

The step (b) oxidation step can be performed using oxidizing agentsincluding oxygen, catalytic reoxidation of palladium,dichlorodicyanoquinone in refluxing xylenes, and palladium acetate inwarm THF. A preferred oxidizing agent is palladijm acetate in warm THF.

Examples 127-129 herein describe a preferred aspect of this method foralkylating a 3-nitro-2-oxo-1,2-dihydropyridyl acetate compound at ringposition 4 comprising

(a) combining the compound with zinc chloride and then adding an alkylgrignard under anhydrous coditions to form a3-nitro-2-oxo-4-alkyl-1,2,3,4-dihydropyridyl acetate intermediate,

(b) contacting the intermediate with palladium acetate in warm THF, and

(c) recovering a 4-alkyl-3-nitro-2-oxo-1,2-dihydropyridyl acetateproduct.

The compound of Example 129 is an intermediate in the synthesis ofcompounds such as those in Example 134.

In this preferred aspect the 3-nitro-2-oxo-1,2-dihydropyridyl acetatecompound is t-butyl[3-nitro-2-oxo-1,2-dihydropyridyl]acetate and the4-alkyl-3-nitro-2-oxo-1,2-dihydropyridyl acetate product is t-butyl[3-nitro-2-oxo-4-(3-phenylpropyl)-1,2-dihydropyridyl]acetate.

A further aspect of the present invention is a method for alkylating a3-nitro-2-oxo-1,2-dihydropyridyl acetate compound at ring position 4comprising

(a) combining the compound with a solution of a zinc salt and an alkylgrignard under anhydrous conditions to form a3-nitro-2-oxo-4-alkyl-1,2,3,4-dihydropyridyl acetate intermediate,

(b) contacting the intermediate with a reducing agent, and

(c) recovering a 4-alkyl-3-amino-2-oxo-piperidyl acetate product.

The disclosed method provides an efficient route to the synthesis of anintermediate used in the synthesis of copounds having a 6-memberedheterocycle comprisng an alkyl at the four position and an amino at thethree position.

Zinc salts such as zinc chloride, zinc bromide, and zinc iodide can beused in step (a). Zinc chloride and zinc bromide are preferred, and zincchloride is especially preferred.

The alkyl grignard in (a) is synthesized fro a starting compound in thegroup defined by [R1]. A preferred alkyl grignard is 3-phenylpropylmagnesium bromide.

Step (a) can be performed by combining the compound and the zinc salt,followed by addition of the alkyl grignard, or by combining the zincsalt and the grignard, followed by addition of the compound. Preferredis a method wherein the compound and the zinc salt are combined, folledby addition of the alkyl grignard.

The step (b) reduction step can be performed with a reducing agent suchas hydrogen, which is preferred.

On aspect of ths method for alkylating a3-nitro-2-oxo-1,2-dihydropyridyl acetate copound at ring position 4comprises

(a) combining the compound with zinc chloride and then adding an alkylgrignard under anhydrous conditions to form a3-nitro-2-oxo-4-alkyl-l,2,3,4-dihydropyridyl acetate intermediate,

(b) contacting the intermediate with hydrogen, and

(c) recovering a 4-alkyl-3-amino-2-oxo-piperidyl acetate product.

3. Selection of Preferred Compounds

The compounds of the present invention are screened for their ability toinhibit some or all of thrombin, factor Xa, plasmin, recombinant tissueplasminogen activator (rt-PA), activated protein C (aPC), chymotrypsin,and trypsin as set forth below. Certain of the preferred compounds aredistinguished by their ability to inhibit thrombin, while notsubstantially inhibiting some or all of factor Xa, plasmin, t-PA, aPC,chymotrypsin, and trypsin. With respect to thrombin and the otherenzymes and as used herein, the term "not substantially inhibiting"means that the IC₅₀ (or K_(i)) for plasmin, t-PA, aPC, chymotrypsin, andtrypsin for a given compound is greater than or equal to its IC₅₀ (orK_(i), respectively) for thrombin.

The compounds of the present invention are dissolved in buffer to givesolutions containing concentrations such that assay concentrations rangefrom 0 to 100 micromolar. In the assays for thrombin, factor Xa,plasmin, t-PA, aPC, chymotrypsin, and trypsin, a chromogenic syntheticsubstrate is added to a solution containing test compound and the enzymeof interest, and the residual catalytic activity of that enzyme isdetermined spectrophometrically. The IC₅₀ of a compound of the presentinvention is determined from the rate of substrate turnover caused bythe specific enzyme being measured. IC₅₀ is that concentration of testcompound giving 50% inhibition of the rate of substrate turnover.Likewise, the K_(i) of a compound of the present invention is determinedfrom the rate of substrate turnover caused by the specific enzyme beingmeasured at various enzyme concentrations. K_(i) is that concentrationof test compound giving 50% inhibition of the rate of substrateturnover. Examples A and B provide an exemplar of the in vitro assaysused to select the compounds of the present invention.

Certain of the preferred compounds of the present invention have a K_(i)of about 0.001 to about 200 nM in the thrombin assay. Especiallypreferred compounds have a K_(i) of about 0.001 to about 50 nM. The moreespecially preferred compounds have a K_(i) of about 0.001 to about 10nM.

Certain of the preferred compounds of the present invention have a IC₅₀for factor Xa, plasmin, t-PA, aPC, chymotrypsin, and trypsin which is atleast 10 times greater than its IC₅₀ for thrombin. Especially preferredcompounds have an IC₅₀ for factor Xa, plasmin, rt-PA, aPC, chymotrypsin,and trypsin which is about 20 to about 100,000 times greater than itsIC₅₀ for thrombin. More especially preferred compounds have an IC₅₀ forfactor Xa, plasmin, rt-PA, aPC, chymotrypsin, and trypsin which is about100 to about 1,000,000 times greater than its IC₅₀ for thrombin. In theevent that a compound of the present invention has an IC₅₀ with respectto factor Xa, plasmin, rt-PA, aPC, chymotrypsin, or trypsin which isgreater than the highest concentration of compound tested, the IC₅₀ istaken to be that highest concentration of compound.

Example B also provides a method for identifying and selecting compoundsof the present invention that inhibit factor Xa, plasmin, t-PA, aPC,chymotrypsin and trypsin to a greater extent than they inhibit thrombinand, thus, have utility as inhibitors of those proteases.

4. Pharmaceutical Compositions

In another aspect, the present invention encompasses pharmaceuticalcompositions prepared for storage or administration which comprise atherapeutically effective amount of a compound of the present inventionin a pharmaceutically acceptable carrier.

The therapeutically effective amount of a compound of the presentinvention will depend on the route of administration, the type of mammalbeing treated, and the physical characteristics of the specific mammalunder consideration. These factors and their relationship to determiningthis amount are well known to skilled practitioners in the medical arts.This amount and the method of administration can be tailored to achieveoptimal efficacy but will depend on such factors as weight, diet,concurrent medication and other factors which those skilled in themedical arts will recognize.

The therapeutically effective amount of the compound of the presentinvention can range broadly depending upon the desired affects and thetherapeutic indication. Typically, dosages will be between about 0.01mg/kg and 100 mg/kg body weight, preferably between about 0.01 and 10mg/kg, body weight.

Pharmaceutically acceptable carriers for therapeutic use are well knownin the pharmaceutical art, and are described, for example, inRemington's Pharmaceutical Sciences, Mack Publishing Co. (A. R. Gennaroedit. 1985). For example, sterile saline and phosphate-buffered salineat physiological pH may be used. Preservatives, stabilizers, dyes andeven flavoring agents may be provided in the pharmaceutical composition.For example, sodium benzoate, sorbic acid and esters of p-hydroxybenzoicacid may be added as preservatives. Id. at 1449. In addition,antioxidants and suspending agents may be used. Id.

The pharmaceutical compositions of the present invention may beformulated and used as tablets, capsules or elixers for oraladministration; suppositories for rectal administration; sterilesolutions and suspensions for injectable administration; and the like.The dose and method of administration can be tailored to achieve optimalefficacy but will depend on such factors as weight, diet, concurrentmedication and other factors which those skilled in the medical artswill recognize.

When administration is to be parenteral, such as intravenous on a dailybasis, injectable pharmaceutical compositions can be prepared inconventional forms, either as liquid solutions or suspensions, solidforms suitable for solution or suspension in liquid prior to injection,or as emulsions. Suitable excipients are, for example, water, saline,dextrose, mannitol, lactose, lecithin, albumin, sodium glutamate,cysteine hydrochloride, or the like. In addition, if desired, theinjectable pharmaceutical compositions may contain minor amounts ofnontoxic auxilliary substances, such as wetting agents, pH bufferingagents, and the like. If desired, absorption enhancing preparations(e.g., liposomes) may be utilized.

5. Utility and Methods

Compounds of the present invention when made and selected as disclosedare useful as potent inhibitors of thrombin in vitro and in vivo. Assuch, these compounds are useful as in vitro diagnostic reagents toprevent the clotting of blood and as in vivo pharmaceutical agents toprevent, inhibit and/or attenuate thrombosis in mammals suspected ofhaving a condition characterized by abnormal thrombosis.

The compounds of the present invention are useful as in vitro diagnosticreagents for inhibiting clotting in blood drawing tubes. The use ofstoppered test tubes having a vaccum therein as a means to draw bloodobtained by venipuncture into the tube is well known in the medicalarts. Kasten, B. L., "Specimen Collection", Laboratory Test Handbook,2nd Edition, Lexi-Comp Inc., Cleveland pp. 16-17 (Edits. Jacobs, D. S.et al. 1990). Such vacuum tubes may be free of clot-inhibitingadditives, in which case, they are useful for the isolation of mammalianserum from the blood. They may alternatively contain clot-inhibitingadditives (such as heparin salts, EDTA salts, citrate salts or oxalatesalts), in which case, they are useful for the isolation of mammalianplasma from the blood. The compounds of the present invention are potentinhibitors of thrombin, and as such, can be incorporated into bloodcollection tubes to prevent clotting of the mammalian blood drawn intothem.

The compounds of the present invention are used alone, in combinationwith other compounds of the present invention, or in combination withother known inhibitors of clotting, in the blood collection tubes. Theamount to be added to such tubes is that amount sufficient to inhibitthe formation of a clot when mammalian blood is drawn into the tube. Theaddition of the compounds to such tubes may be accomplished by methodswell known in the art, such as by introduction of a liquid compositionthereof, as a solid composition thereof, or liquid composition which islyophilized to a solid. The compounds of the present invention are addedto blood collection tubes in such amounts that, when combined with 2 to10 mL of mammalian blood, the concentration of such compounds will besufficient to inhibit clot formation. Typically, the requiredconcentration will be about 1 to 10,000 nM, with 10 to 1000 nM beingpreferred.

The compounds of the present invention are useful as a pharmaceuticalagent for preventing, inhibiting and/or attenuating thrombosis in amammal suspected of having a condition characterized by abnormalthrombosis.

Conditions characterized by abnormal thrombosis are well known in themedical arts and include those involving the arterial and venousvasculature of mammals. With respect to the coronary arterialvasculature, abnormal thrombosis (thrombus formation) characterizes therupture of an established atherosclerotic plaque which is the majorcause of acute myocardial infarction and unstable angina, as well asalso characterizing the occlusive coronary thrombus formation resultingfrom either thrombolytic therapy or percutaneous transluminal coronaryangioplasty (PTCA). With respect to the venous vasculature, abnormalthrombosis characterizes the condition observed in patients undergoingmajor surgery in the lower extremities or the abdominal area who oftensuffer from thrombus formation in the venous vasculature resulting inreduced blood flow to the affected extremity and a predisposition topulmonary embolism. Abnormal thrombosis further characterizesdisseminated intravascular coagulopathy which commonly occurs withinboth vascular systems during septic shock, certain viral infections andcancer, a condition wherein there is rapid consumption of coagulationfactors and systemic coagulation which results in the formation oflife-threatening thrombi occurring throughout the microvasculatureleading to widespread organ failure.

The present invention includes methods for preventing a condition in amammal suspected of having a condition characterized by abnormalthrombosis, comprising administering to said mammal a therapeuticallyeffective amount of a compound or a pharmaceutical composition of thepresent invention.

The compounds or pharmaceutical compositions of the present inventionare administered in vivo, ordinarily in a mammal, preferably in a human.In employing them in vivo, the compounds or pharmaceutical compositionscan be administered to a mammal in a variety of ways, including orally,parenterally, intravenously, subcutaneously, intramuscularly,colonically, rectally, nasally or intraperitoneally, employing a varietyof dosage forms. Administration is preferably parenteral, such asintravenous on a daily basis. Alternatively, administration ispreferably oral, such as by tablets capsules or elixers taken on a dailybasis.

In practicing the methods of the present invention, the compounds orpharmaceutical compositions of the present invention are administeredalone or in combination with one another, or in combination with othertherapeutic or in vivo diagnostic agents.

As is apparent to one skilled in the medical art, a "therapeuticallyeffective amount" of the compounds or pharmaceutical compositions of thepresent invention will vary depending upon the age, weight and mammalianspecies treated, the particular compounds employed, the particular modeof administration and the desired affects and the therapeuticindication. Because these factors and their relationship to determiningthis amount are well known in the medical arts, the determination oftherapeutically effective dosage levels, the amount necessary to achievethe desired result of preventing thrombosis, will be within the ambit ofone skilled in these arts. Typically, administration of the compounds orpharmaceutical composition of the present invention is commenced atlower dosage levels, with dosage levels being increased until thedesired effect of preventing in vivo thrombosis is achieved which woulddefine a therapeutically effective amount. For the compounds of thepresent invention, alone or as part of a pharmaceutical composition,such doses are between about 0.01 mg/kg and 100 mg/kg body weight,preferably between about 0.01 and 10 mg/kg, body weight.

To assist in understanding, the present invention will now be furtherillustrated by the following examples. These examples as they relate tothis invention should not, of course, be construed as specificallylimiting the invention and such variations of the invention, now knownor later developed, which would be within the purview of one skilled inthe art are considered to fall within the scope of the invention asdescribed herein and hereinafter claimed.

EXAMPLES Example 1 Preparation of N-alpha-t-butoxycarbonyl-N^(g)-nitro-L-arginine lactam ##STR27##

N-alpha-t-butoxycarbonyl-Ng-nitroarginine (2.00 g, 6.3 mmole) wasdissolved in tetrahydrofuran (100 mL) by heating the solution to 50° C.The solution was allowed to cool to room temperature. N-methylpiperidine (0.84 mL, 6.9 mmole) was added, and the solution was cooledin an ice bath. Isobutylchloroformate (0.83 mL, 6.3 mmole) was added,and the reaction mixture was stirred at 0° C. for 6 hours. The reactionmixture was stirred for 18 hours while the ice in the Dewar was allowedto melt overnight. The solvent was removed under vacuum. The crudeproduct was dissolved in 20% ethyl acetate/dichloromethane (10 mL), andwas purified by flash chromatography through a 3×5 cm column of silicagel using 20% ethyl acetate/dichloromethane as eluent. 125 mL of eluentwas collected. The solvent was removed under vacuum to afford 1.39 g(74% crude yield) of the title compound as a white foam. R_(f) =0.44(silica gel, 5% isopropanol in dichloromethane). Isobutanol was presentas an impurity. This compound may be further purified byrecrystallization from dichloromethane/hexanes or ethanol/water.

Example 2 Preparation of N-alpha-t-butoxycarbonyl-N^(g)-nitro-L-argininal ##STR28## (a) Procedure 1

To a stirred solution of LiAlH₄ in tetrahydrofuran (3.8 mL of a 1.0Msolution, 3.8 mmole), cooled in an ice bath, was added dropwise ethylacetate (0.43 mL, 3.8 mmole) in tetrahydrofuran (5 mL). The solution wasstirred for 30 minutes at 0° C. to preform LiAlH₂ (OEt)₂.

The solution of this LiAlH₂ (OEt)₂ was added dropwise to a stirredsolution of compound of Example 1 (0.92 g, 3.1 mmole) in tetrahydrofuran(5 mL). After 30 minutes, the reaction is quenched with 1.0NHCl/tetrahydrofuran (2 mL of a 1:1 mixture). 1.0N HCl (20 mL) was added,and the solution was extracted three times with ethyl acetate (20 mLeach). The combined organic layers were washed with water (5 mL),saturated sodium bicarbonate (5 mL) and twice with brine(5 mL each),dried over anhydrous magnesium sulfate, filtered and the solvent wasremoved under vacuum to give 0.94 g (100% yield) of the title compoundas an off-white solid.

(b) Procedure 2

Alternatively, the title compound was made by the procedures whichfollow.

A 12 liter four-necked round bottom flask equipped with an overheadstirring apparatus was flame dried under a strong stream of nitrogen.After the flask had cooled, 120.0 g of N-alpha-t-butoxycarbonyl-N^(g)-nitro-L-arginine (376 mmole, 1 equivalent) was added under a blanket ofnitrogen followed by the addition of 6 liters of anhydroustetrahydrofuran (Aldrich sure-seal) via canula. The flask was thenfitted with a thermometer and the resulting suspension was warmed to 50°C. with a heat gun while stirring. The reaction mixture was cooled to 5°C. with an ice bath and further cooled to -5° C. with an ice/acetonebath.

During the time it took for this solution to reach -5° C., 36.66 g ofN-methyl-O-methylhydroxyamine hydrochloride (376 mmole, 1.0 equivalent)was weighed out in a 500 mL flask and suspended in 300 mL ofdichloromethane. This suspension was sparged with nitrogen for 5minutes, cooled to 0° C. and 46 mL of N-methylpiperidine (1.0equivalent) was added via syringe under nitrogen. The mixture wassonicated briefly to insure complete dissolution/free base formation andrecooled to 0° C. in an ice bath while still under nitrogen. Theresulting solution of free base was used later.

When the above arginine solution had reached -5° C., 45 mL ofN-methylpiperidine was added via syringe followed 5 minutes later by theaddition of 46 mL of isobutyl chloroformate (0.95 equivalent) viasyringe. The resulting solution was stirred for 15 minutes at -5° C.After this time, the free base solution of N-methyl-O-methylhydroxylamine generated above was added via canula over about 15minutes. Stirring was continued at -5° C. for another 1.5 hours at whichtime thin layer chromatography (silica gel, 1:10:90 aceticacid/methanol/dichloromethane) indicated that the reaction was complete.The reaction mixture was filtered while still cold, the salts washedwith 400 mL of cold tetrahydrofuran and the filtrate concentrated undervacuum on a rotary evaporator to yield a yellow foam.

The crude intermediate was taken up in 300 mL of dichloromethane andapplied to a column of silica gel (70-230 mesh, 7×50 cm). The column wasfirst eluted with 2 liters of dichloromethane followed by 2 liters of 2%methanol in dichloromethane. This was followed by elution with 5%methanol in dichloromethane until all of the product had been eluted(the eluant was checked for UV activity and five one-liter fractionswere collected once this UV activity was apparent). Fractions containingpure product were pooled and concentrated under vacuum and pumped onovernight to yield 120.1 g (88% yield) of N-alpha-t-butoxycarbonyl-N^(g)-nitro-L-arginine-(N-methyl, N-methoxyamide) as light yellow foam. Thisfoam was taken up in 300 mL of dichloromethane, 300 mL of toluene, andthe volatiles were once again removed under vacuum to remove anyresidual water or methanol.

120.1 g of N-alpha-t-butoxycarbonyl-N^(g) -nitro-L-arginine-(N-methyl,N-methoxyamide) (331.4 mmole) was taken up in 2.8 liters of dry (Aldrichsure-seal) tetrahydrofuran and transferred to a dry 5 liter 4-neckedround bottom flask equipped with a mechanical stirrer and a lowtemperature thermometer. The solution was cooled to -70° C. with a dryice/acetone bath and 300 mL of 1M LiAlH₄ in tetrahydrofuran was added bycanula transfer directly from 100 mL Aldrich sure-seal bottles. Anadditional 50 mL of 1M LiAlH₄ in tetrahydrofuran was added via syringe(total 331 mL). During the additions, the reaction temperature was keptbelow -60° C. The reaction was stirred for 0.5 hours at -70° C., thecooling bath removed, and the reaction was slowly allowed to warm to 0°C. (about 2.5 hours). Between -30° C. and -20° C. a thick slurryresulted. When the reaction mixture obtained 0° C., a small aliquot wasremoved and partitioned between ethyl acetate/2M potassium bisulfate.The organic layer was analyzed by thin layer chromatography (silica gel,ethyl acetate).

When the reaction was judged to be complete, it was cooled to -70° C.and 503 mL of 2M potassium bisulfate was added via dropping funnel at aslow enough rate to keep the reaction temperature below -30° C. Thecooling bath was removed and the reaction mixture was allowed to come to0° C. over the course of 2 hours at which time a white precipitate wasfiltered off. The solids were washed with 500 mL of coldtetrahydrofuran. The filtrate was concentrated under vacuum on a rotaryevaporator until most of the tetrahydrofuran was removed and theremaining white sludge was mostly aqueous. The crude product was takenup in 1.5 liters of ethyl acetate and washed with 0.2 M HCl (2×200 mL).The HCl extracts were back-extracted with 400 mL of ethyl acetate andthe organics were combined and extracted with saturated sodiumbicarbonate (2×200 mL). The bicarbonate extracts were alsoback-extracted with 400 ml of ethyl acetate. The organics were thencombined and washed with brine (200 mL) followed by drying overanhydrous sodium sulfate. The solution was filtered, concentrated undervacuum on a rotary evaporator and pumped on overnight to yield a whitesolid (89.0 g) of crude title compound. This was chromatographed onsilica gel and eluted with a gradient of 0 to 10% methanol indichloromethane. The pure fractions were combined and evaporated toyield the title compound as a white solid (75 g, 74%).

Example 3 Preparation of N-alpha-t-butoxycarbonyl-N^(g)-nitro-L-argininal ethyl cyclol ##STR29##

The compound of Example 2 (41.60 g, 0.137 mole) was dissolved in ethanol(200 mL) and concentrated HCl (1 mL) was added. After the reaction wascomplete by TLC (silica gel, 10% methanol in dichloromethane), thesolvent was removed under vacuum. The crude product was purified byflash chromatography through a column of silica gel (230-400 mesh) using0 to 10% ethyl acetate/dichloromethane as eluent. The combined fractionsyielded 36.88 g (81%) of the title compound as pale yellow foam. R_(f)=0.62 (silica gel, 5% methanol in dichloromethane).

Example 4 Preparation of N^(g) -nitro-L-argininal ethyl cyclol,hydrochloride salt ##STR30##

To a solution of the compound of Example 3 (35 g) in 500 mL of absoluteethanol at 0° C. was added slowly 500 mL of absolute ethanol saturatedwith HCl(g). This mixture was allowed to warm to 25° C. and checked bythin-layer chromatography. The appearance of a very polar product wasthe desired compound. Most of the HCl was removed with a stream of drynitrogen and the resulting organic solvent was removed under vacuum. Theresulting 33 g of the title compound as a yellow-white solid was usedwithout further purification.

Example 5 Preparation of ethyl (3-nitro-2-oxo-1,2-dihydropyridyl)acetate##STR31##

Sodium hydride (4.08 g of a 60% dispersion in mineral oil, 0.10 mole)was washed with hexanes three times (10 mL each) and suspended indimethylformamide (50 mL). The stirred suspension was cooled in an icebath, then 3-nitro-2-hydroxypyridine (13.0 g, 0.093 mole) was added insmall portions over a 45-minute period. After the addition was complete,the reaction was stirred at 0° C. for 10 minutes, then room temperaturefor 30 minutes. The reaction mixture was recooled in an ice bath. Ethylbromoacetate (0.75 mL, 0.097 mole) was added. The reaction was stirredat 0° C. for 1 hour, then 1.5 hours at room temperature. The reactionmixture was partitioned between ethyl acetate (200 mL) and water (200mL). The aqueous layer was extracted with ethyl acetate (3×200 mL). Thecombined organic extracts were washed with water (4×100 mL), brine, anddried over anhydrous sodium sulfate. The solvent was removed underreduced pressure. The residue was chromatographed through silica gelusing 0 to 20% ethyl acetate/dichloromethane as eluent to afford 15.7 g(75% yield) of the title compound as yellow solid. R_(f) =0.30 (silicagel, 20% ethyl acetate in dichloromethane).

Example 6 Preparation of ethyl(3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl)acetate ##STR32##

A stirred solution of the compound of Example 5 (44.5 g, 0.197 mole) inethanol (200 mL) was hydrogenated over 10% Pd/C (2.25 g) for 16 hoursunder balloon pressure. Celite was added, and the reaction mixture wasfiltered through a pad of celite in a 600 mL fritted funnel (5 cmdepth), using ethyl acetate to wash. The solvent was removed undervacuum, diluted with ethyl acetate (200 mL) and toluene (200 mL), andthe solvent was removed under vacuum to give crude ethyl(3-amino-2-oxo-1,2-dihydropyridyl)acetate (40.0 g, 0.204 mole) inquantitative yield.

A stirred solution of ethyl (3-amino-2-oxo-1,2-dihydropyridyl)acetate(40.0 g, 0.204 mole) and 2,4,6-collidine (54 mL, 0.408 mole) intetrahydrofuran (200 mL) was cooled in an ice bath. A solution ofbenzylsulfonyl chloride (38.9 g, 0.204 mmole) in tetrahydrofuran (200mL) was added over a 50-minute period. After addition was complete, thesolution was stirred for 30 minutes at 0° C. The reaction mixture wasdiluted with ethyl acetate (1.2 L), washed with 1.0N HCl (until aqueouslayer is pH 1), water (50 mL), saturated sodium bicarbonate (100 mL),and brine (2×50 mL). The organic layer was dried over anhydrousmagnesium sulfate, and the solvent was removed. The residue wasrecrystallized from chloroform. 39 g of the title compound was isolated.To the mother liquor was added silica gel. The solution was swirled,then filtered through a sintered glass funnel, washing with 50% ethylacetate in dichloromethane. The solvent was removed from the filtrate,and the residue was recrystallized from chloroform. An additional 13 gof the title compound was isolated to afford a total of 52.00 g (75%yield) of the title compound as a tan solid. R_(f) =0.32 (silica gel,20% ethyl acetate in dichloromethane); m.p. 48-49° C.

Example 7 Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetic acid##STR33##

To a cooled (0° C.) suspension of the compound of Example 6 (50.89 g,0.145 mole) in methanol (500 mL) was added 1.0N NaOH (327 mL) over aperiod of 10 minutes. After the addition was complete, the solution wasallowed to warm to room temperature over a period of 1.5 hours. Thesolution became homogeneous upon addition of NaOH. A precipitate formedduring the reaction. The solvent was reduced under vacuum, the residuediluted with water (400 mL), and washed with ethyl acetate (2×150 mL).The aqueous layer was acidified with 2.0N HCl to pH 1, and extractedwith ethyl acetate (3×200 mL). The combined organic extracts were washedwith water, then brine (twice). The product crystallized. The 2 combinedcrops yielded 44.54 g (95%) of the title compound as off-white crystals.R_(f) =0.17 (silica gel, 1% acetic acid, 10% methanol indichloromethane); m.p. 186-187° C.

Example 8 Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-N^(g)-nitro-L-argininal ethyl cyclol ##STR34##

To a stirred suspension of the compound of Example 7 (23.3 g, 77 mmole),the compound of Example 4 (N^(g) -nitro-L-argininal ethyl cyclol,hydrochloride salt, 24.76 g, 92 mmole), and N-hydroxybenzotriazole(11.79 g, 77 mmole) in acetonitrile (400 mL) cooled to 0° C. was added1-ethyl-3-(3-dimethylamino-propyl)carbodiimide hydrochloride salt (EDC,17.76 g, 92 mmole). After 30 minutes, the solution was almosthomogeneous. N-methylmorpholine (25.4 mL, 231 mmole) was added dropwise.After the addition was complete, the reaction was stirred at roomtemperature for 3 hours. The solvent was reduced under vacuum, and theresulting residue was dissolved in dichloromethane (600 mL), washed with50 mL each of 2.0N HCl (to pH 1), water, saturated sodium bicarbonateand brine. The extract was dried over anhydrous magnesium sulfate, andthe solvent was removed under vacuum. The crude product was purified inthree batches using a 600 mL fritted funnel as a column through silicagel (7 cm depth) to yield (29.40 g, 74%) of the title compound.Analytical HPLC gave t_(R) =12.8 minutes (20-60% CH₃ CN, 25 mm VydacC-18 column). R_(f) =0.28 (silica gel, 5% ethanol in dichloromethane).

Example 9 Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalethyl cyclol, acetate salt ##STR35##

The compound of Example 8 (5.60 g, 11 mmole) in ethanol/aceticacid/water (4:1:1, 60 mL) was hydrogenated over 10% palladium on carbon(1.80 g) for 4 hours at 20 psi. Celite was added, and the solution wasfiltered through a 0.2 micron filter, washing the solid withethanol/acetic acid/water (4:1:1, 60 mL). To the filtrate was added 10%palladium on carbon (1.80 g), and the solution was hydrogenated at 20 to25 psi for 40 hours. Celite was added, and the solution was filteredthrough a 0.2 micron filter, washing the solid with water (200 mL). Thesolvent was reduced to a volume of 200 mL under reduced pressure, thenwashed with ethyl acetate (50 mL). The solvent from the aqueous layerwas reduced to remove the volatiles, then the aqueous layer waslyophilized to yield 4.88 g (85% yield) of the title compound.Analytical HPLC gave t_(R) =9.5 minutes (20-60% CH₃ CN, 25 mm Vydac C-18column).

Example 10 Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,trifluoroacetate salt ##STR36##

The compound of Example 9 (4.88g, 9.2 mmole) was suspended in 3.0 N HCl(100 mL). After stirring for 3 h, the reaction mixture was quenched with2.5 M aqueous sodium acetate to pH 3.5 to 4, then filtered through a 2micron filter. The filtrate was purified in two batches by preparativeHPLC (Waters PrepPak cartridge, Delta-Pak C18, 300 angstrom column, 0 to40% acetonitrile/water containing 0.1% trifluoroacetic acid). The cleanfractions were combined to give 2.05 g (40% yield) of the titlecompound. Fast atom bombardment mass spectrometry confirmed thetheoretical molecular weight of 463.

Example 11 Preparation of 6-methylpyrid-2-one-3-carbonitrile ##STR37##

To stirred a mixture of sodium methoxide (46.5 g, 860 mmole) in diethylether (950 mL), cooled in an ice bath, was added a mixture of acetone(46.5 g, 800 mmole) and ethyl formate (59.6 g, 800 mmole) dropwise over1 hour. When addition was complete, the cooling bath was removed and themixture was warmed to room temperature over a 1 hour period. Thevolatile materials were distilled, keeping the oil bath at no more than60° C. To the solid residue was added cyanoacetamide (67 g, 800 mmole)in water (400 mL) and piperidine acetate (140 mmole, prepared by addingpiperidine to a solution of 8.0 mL of acetic acid in 20 mL water untilthe solution was greater than pH 7). The flask was fitted with a refluxcondenser, and the mixture was heated for 2 hours under reflux. Themixture was cooled to room temperature and acidified to pH 5 with aceticacid. After standing overnight at room temperature, the mixture wascooled in an ice bath for 45 minutes. The yellow solid product wasfiltered, washed with ice water four times and dried under vacuum at 80°C. overnight. Crystallization from 50% (v/v) ethanol afforded the titlecompound as a yellow solid (52.6 g, 49% yield). R_(f) =0.29 (silica gel,95:5 chloroform:methanol). Fast atom bombardment mass spectrometryconfirmed the theoretical molecular weight of 134.

Example 12 Preparation of 6-methylpyrid-2-one-3-carboxylic acid##STR38##

A suspension of the compound of example 11 (16.9 g) in 20% NaOH (w/w, 63mL) was heated at 140 to 145° C. overnight in a sealed bomb. The cooledreaction mixture was acidified to about pH 8 with concentratedhydrochloric acid and extracted with dichloromethane (three times). Theaqueous phase was acidified, precipitating a yellow solid which wasfiltered, washed with water, and dried overnight in a vacuum oven atapproximately 80° C. The dried title compound (15.68 g, 81% yield)required no further purification. Fast atom bombardment massspectrometry confirmed the theoretical molecular weight of 153.

Example 13 Preparation of 3-benzyloxycarbonylamino-6-methylpyrid-2-one##STR39##

To the compound of Example 12 (11.8 g, 0.077 mole), suspended in dioxane(260 mL), triethylamine (11.3 mL, 0.081 mole) is added dropwise rapidlywith stirring followed by diphenylphosphoryl azide (16.7 mL, 0.077mole). The suspension is heated under reflux for 4 hours using apreheated 120° C. oil bath. Benzyl alcohol (24.1 mL, 0.23 mole) is thenadded and the mixture was stirred under reflux overnight. The reactionmixture is cooled and evaporated. The residue is suspended in water (600mL) and filtered. The filter cake is washed with 10% HCl twice,saturated sodium bicarbonate and brine. The crude product ischromatographed using 20 to 30% ethyl acetate/chloroform to give thetitle compound.

Example 14 Preparation of ethyl(3-benzyloxycarbonylamino-6-methyl-2-oxo-1,2-dihydro-1-pyridyl)acetate##STR40##

The compound of Example 13 (1.80 g, 7.0 mmole) was added to a stirredsuspension of sodium hydride (0.33 g, 8.4 mmole) in drydimethylformamide (50 mL). After 45 minutes, ethyl iodoacetate (1.43 g,6.7 mmole) was added, and the mixture was stirred overnight, dilutedwith 10% hydrochloric acid (300 mL) and extracted with ethyl acetate(3×150 mL). The organic layer was washed with brine (twice), dried andevaporated. The resulting yellow, waxy solid was chromatographed,eluting with 3% ethyl acetate in dichloromethane, to give the titlecompound (1.28 g, 53% yield). R_(f) =0.52 (silica gel, 5:95methanol:dichloromethane). Fast atom bombardment mass spectrometryconfirmed the theoretical molecular weight of 344.

Example 15 Preparation of ethyl[3-[(benzylsulfonyl)amino]-6-methyl-2-oxo-1,2-dihydropyridyl]acetate##STR41##

A stirred solution of compound of Example 14 (2.50 g) in ethanol (25 mL)is hydrogenated over 10% Pd/C (0.25 g) for 5 hours under balloonpressure. Celite is added, and the reaction mixture is filtered througha pad of celite, using ethyl acetate to wash. The solvent is removedunder vacuum, diluted with ethyl acetate (20 mL) and toluene (20 mL),and the solvent is removed under vacuum to give crude ethyl(3-amino-6-methyl-2-oxo-1,2-dihydropyridyl)acetate.

A stirred solution of ethyl(3-amino-6-methyl-2-oxo-1,2-dihydropyridyl)acetate (0.55 g, 2.6 mmole)and 2,4,6-collidine (1.2 mL, 5.2 mmole) in tetrahydrofuran (10 mL) iscooled in an ice bath. A solution of benzylsulfonyl chloride (0.50 g,2.6 mmole) in tetrahydrofuran (10 mL) is added over a 15 minute period.After addition is complete, the solution is stirred for 30 minutes at 0°C. The reaction mixture is diluted with ethyl acetate (100 mL), washedwith 1.0N HCl (until aqueous layer is pH 1), water (10 mL), saturatedsodium bicarbonate (10 mL), and brine (2×10 mL). The organic layer isdried over anhydrous magnesium sulfate, and the solvent is removed. Theresidue is chromatographed through silica gel using 0:100-20:80 ethylacetate:dichloromethane as eluent to afford the title compound.

Example 16 Preparation of[3-[(benzylsulfonyl)amino]-6-methyl-2-oxo-1,2-dihydropyridyl]acetic acid##STR42##

Using similar procedures to that described above in Example 7, the titlecompound is prepared from the compound of Example 15. An alternativemethod of preparing the title compound is described in Examples 91 to96.

Example 17 Preparation of[3-[(benzylsulfonyl)amino]-6-methyl-2-oxo-1,2-dihydropyridyl]acetyl-N.sup.g-nitro-L-argininal ethyl cyclol ##STR43##

Using similar procedures to that described above in Example 8, the titlecompound is prepared from the compound of Example 16.

Example 18 Preparation of[3-[(benzylsulfonyl)amino]-6-methyl-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalethyl cyclol, acetate salt ##STR44##

Using similar procedures to that described above in Example 9, the titlecompound is prepared from the compound of Example 17.

Example 19 Preparation of[3-[(benzylsulfonyl)amino]-6-methyl-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,trifluoroacetate salt ##STR45##

Using similar procedures to that described above in Example 10, thetitle compound is prepared from the compound of Example 18. Analternative method of preparing the title compound is described inExample 113 (Compound C).

Example 20 Preparation of ethyl pyrimidin-6(1H)-one-5-carboxylate##STR46##

Diethyl ethoxymethylenemalonate (10.1 mL, 50 mmole) and formamidineacetate (10.4 g, 100 mmole) were refluxed in ethanol (10 mL) for 24hours. The reaction mixture was allowed to cool to room temperatureovernight, and suspended in ethyl acetate (30 mL) and 1.0N HCl (20 mL).The suspension was filtered, and the filter cake was washed with 1.0NHCl, followed by water, then ethyl acetate, and air dried affording thetitle compound as an tan solid (3.33 g, 40% yield). R_(f) =0.21 (silicagel, 10% methanol in dichloromethane); m.p. 187-188° C.

Example 21 Preparation of ethyl1-allyl-pyrimidin-6(1H)-one-5-carboxylate ##STR47##

The compound of Example 20 (4.3 g, 26 mmole) is added to a stirredsuspension of sodium hydride (1.13 g, 28 mmole) in dry dimethylformamide(50 mL). After 45 minutes, allyl bromide (2.21 mL, 26 mmole) is added,and the mixture is stirred overnight, diluted with 10% hydrochloric acid(300 mL) and extracted with ethyl acetate (3×150 mL). The organic layeris washed with brine (twice), dried and evaporated. The residue ischromatographed through silica gel using 0-10%isopropanol/dichloromethane as eluent. The title compound is isolated.

Example 22 Preparation of 1-allyl-pyrimidin-6(1H)-one-5-carboxylic acid##STR48##

To the compound of Example 21 (5.00 g, 0.024 mole), suspended inmethanol (25 mL) and cooled in an ice bath, 1.0N NaOH (29 mL, 0.029mole) is added dropwise rapidly with stirring. After 16 hours, thesolvent is reduced under vacuum, residue diluted with water (50 mL), andwashed with ethyl acetate (2×15 mL). The aqueous layer is acidified with2.0N HCl to pH 1, extracted with ethyl acetate (3×50 mL). The combinedorganic extracts are washed with water, then brine (twice). The solventis removed in vacuo to afford the title compound.

Example 23 Preparation of1-allyl-5-t-butyloxycarbonylamino-6-oxo-1-pyrimidi-6(1H)-one ##STR49##

To the compound of Example 22 (10.0 g, 0.056 mole), suspended in dioxane(260 mL), triethylamine (8.1 mL, 0.058 mole) is added dropwise rapidlywith stirring followed by diphenylphosphoryl azide (12.0 mL, 0.077mole). The suspension is heated under reflux for 4 hours using apreheated 120° C. oil bath. t-Butanol (12.3 g, 0.17 mole) is then addedand the mixture is stirred under reflux overnight. The reaction mixtureis cooled and evaporated. The residue is suspended in water (600 mL) andfiltered. The filter cake is washed with 1.0N HCl twice, saturatedsodium bicarbonate and brine. The crude product is chromatographed using0 to 50% ethyl acetate/dichloromethane to give the title compound.

Example 24 Preparation of 1-allyl-5-amino-6-oxo-1-pyrimidi-6(1H)-one,trifluoroacetate salt ##STR50##

The compound of Example 23 (5.00 g) is treated with 50% trifluoroaceticacid in dichloromethane (50 mL) for 35 minutes. The solution is addeddropwise to diethyl ether (500 mL), while swirling. The precipitate isfiltered, washing with diethyl ether. The powder is dried under vacuumto yield the title compound.

Example 25 Preparation of1-allyl-5-benzylsulfonylamino-6-oxo-1-pyrimidi-6(1H)-one ##STR51##

The free base of the compound of Example 24 is generated by dissolvingthe compound of Example 24 (5.00 g, 18.9 mmole) in 1M potassiumcarbonate. The free base is then extracted into dichloromethane, whichis dried and evaporated to give1-allyl-5-amino-6-oxo-1-pyrimidi-6(1H)-one.

A stirred solution of 1-allyl-5-amino-6-oxo-1-pyrimidi-6(1H)-one and2,4,6-collidine (8.3 mL, 38 mmole) in tetrahydrofuran (25 mL) is cooledin an ice bath. A solution of benzylsulfonyl chloride (3.59 g, 18.9mmole) in tetrahydrofuran (25 mL) is added over a 15-minute period.After addition is complete, the solution is stirred for 1 hour at 0° C.The reaction mixture is diluted with ethyl acetate (200 mL), washed with1.0N HCl (until aqueous layer is pH 1), water (25 mL), saturated sodiumbicarbonate (25 mL), and brine (2×25 mL). The organic layer is driedover anhydrous magnesium sulfate and the solvent is removed in vacuo togive the title compound.

Example 26 Preparation of5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetaldehyde##STR52##

To a solution of the compound of Example 25 (5.00 g, 16.4 mmole) intetrahydrofuran (50 mL) and water (7 mL) is addedN-methylmorpholine-N-oxide (3.20 g, 16.4 mmole) and osmium tetroxide(1.0 mL of a 4% solution in water, 0.16 mmole). After the reactionmixture is stirred for 18 hours, N-methylmorpholine-N-oxide (0.47 g, 2.8mmole) is added. After stirring the reaction mixture for 4 hours, sodiumthiosulfate, (2.5 mL of a saturated aqueous solution) and diatomaceousearth (7 g) are added, and the mixture is stirred for 30 minutes. Themixture is filtered and evaporated under vacuum to give an oil. This oilis dissolved in ethanol (60 mL) and a solution of sodium periodate (7.00g, 33 mmole) in water (10 mL) is added. The residue is dissolved inethyl acetate and the solution is washed with water, dried, andevaporated to afford the title compound.

Example 27 Preparation of5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetic acid##STR53##

To the compound of Example 26 (5.00 g, 15.6 mmole) in t-butanol (40 mL)and 2-methyl-2-butene (33 mL) is added a solution of sodium chlorite (13g, 14 mmole) and sodium dihydrogen phosphate monohydrate (15.1 g, 109mmole) in water (40 mL). The reaction mixture is stirred for 3 hours,then concentrated under reduced pressure. The residue is diluted withethyl acetate and extracted with 1.0N sodium hydroxide. The aqueouslayer is acidified to pH 1 with 1.0N hydrochloric acid, then extractedwith dichloromethane twice. The organic extracts are dried andevaporated to give the title compound.

Example 28 Preparation of5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetyl-N^(g)-nitro-L-argininal ethyl cyclol ##STR54##

Using similar procedures to that described above in Example 8, the titlecompound is prepared from the compound of Example 27.

Example 29a Preparation of5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetyl-L-argininalethyl cyclol, acetate salt ##STR55##

Using similar procedures to that described above in Example 9, the titlecompound is prepared from the compound of Example 28.

Example 29b Preparation of5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetyl-L-argininal,trifluoroacetate salt ##STR56##

Using similar procedures to that described above in Example 10, thetitle compound is prepared from the compound of Example 29a.

Example 30 Ethyl 2-methyl-pyrimidin-6(1H)-one-5-carboxylate ##STR57##

Acetamidine hydrochloride (37.16 g, 0.39 mole) was stirred in sodiumethoxide in ethanol (73 mL of a 21% solution, 0.20 mole) for 5 minutes.Diethyl ethoxymethylenemalonate (31.5 mL, 0.15 mole) was added, and thereaction mixture was refluxed for 5 hours. The reaction mixture wasallowed to cool to room temperature overnight, and diluted withdichloromethane (100 mL). The solution was filtered, washing the solidcake with dichloromethane. The filtrate was concentrated at reducedpressure. The residue was dissolved in dichloromethane (150 mL) and 2.0NHCl (30 mL). The pH of the aqueous layer was 1. The organic layer waswashed with water, saturated sodium bicarbonate and brine, dried overanhydrous magnesium sulfate, and the solvent was removed under reducedpressure. The residue was dissolved in hot dichloromethane (50 mL).Ethyl acetate was added (50 mL). The product precipitated. The solutionwas boiled for 5 minutes, cooled to room temperature, and hexanes wereadded (50 mL). The resulting crystals were filtered, then washed withethyl acetate (20 mL) followed by hexanes (50 mL) to yield the titlecompound (7.22 g, 27%) as off-white crystals. R_(f) =0.27 (silica gel,10% isopropanol in dichloromethane). The title compound also wasprepared by the route described in Example 102.

Example 31 Preparation of ethyl1-allyl-2-methyl-pyrimidin-6(1H)-one-5-carboxylate ##STR58##

Using similar procedures to that described above in Example 21, thetitle compound is prepared from the compound of Example 30.

Example 32

Preparation of 1-allyl-2-methyl-pyrimidin-6(1H)-one-5-carboxylic acid##STR59##

Using similar procedures to that described above in Example 22, thetitle compound is prepared from the compound of Example 31.

Example 33 Preparation of1-allyl-2-methyl-5-t-butyloxycarbonylamino-6-oxo-1-pyrimidi-6(1H)-one##STR60##

Using similar procedures to that described above in Example 23 the titlecompound is prepared from the compound of Example 32.

Example 34 Preparation of1-allyl-2-methyl-5-amino-6-oxo-1-pyrimidi-6(1H)-one, trifluoroacetatesalt ##STR61##

Using similar procedures to that described above in Example 24, thetitle compound is prepared from the compound of Example 33.

Example 35 Preparation of1-allyl-2-methyl-5-benzylsulfonylamino-6-oxo-1-pyrimidi-6(1H)-one##STR62##

Using similar procedures to that described above in Example 25, thetitle compound is prepared from the compound of Example 34.

Example 36 Preparation of5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetaldehyde##STR63##

Using similar procedures to that described above in Example 26, thetitle compound is prepared from the compound of Example 35.

Example 37 Preparation of5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetic acid##STR64##

Using similar procedures to that described above in Example 27, thetitle compound is prepared from the compound of Example 36. Analternative method of preparing the title compound is described inExamples 102 to 107.

Example 38 Preparation of2-methyl-5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetyl-N.sup.g-nitro-L-argininal ethyl cyclol ##STR65##

Using similar procedures to that described above in Example 8, the titlecompound is prepared from the compound of Example 37.

Example 39 Preparation of2-methyl-5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetyl-L-argininalethyl cyclol, acetate salt ##STR66##

Using similar procedures to that described above in Example 9, the titlecompound is prepared from the compound of Example 38.

Example 40 Preparation of2-methyl-5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetyl-L-argininal,trifluoroacetate salt ##STR67##

Using similar procedures to that described above in Example 10, thetitle compound is prepared from the compound of Example 39. Analternative method of preparing the title compound is described inExample 113 (Compound D).

Example 41 Preparation of 5-nitro-1-methoxymethyluracil ##STR68##

5-nitrouracil (10.00 g, 64 mmole), 1,1,1-3,3,3-hexamethyldisilazane (40ML, 190 mmole) and chlorotrimethylsilane (4.0 mL, 32 mmole) are heatedto reflux for 24 hours. The solution is concentrated under reducedpressure to afford 5-nitrouracil bis(trimethylsilyl) ether.5-nitrouracil bis(trimethylsilyl) ether (10.0 g, 24 mmole),dimethylformamide (50 mL) and bromomethylmethyl ether (5.9 mL, 73 mmole)are heated in an 80° C. oil bath for 24 hours. Ice water (500 mL) isadded, and the mixture is stirred for 30 minutes, then extracted withdichloromethane (3X). The combined organic layers are dried overanhydrous magnesium sulfate, filtered, and concentrated under reducedpressure to give the title compound.

Example 42 Preparation of ethyl5-nitro-1-methoxymethyl-3-uracilylacetate ##STR69##

The compound of Example 41 (10.0 g, 50 mmole) is dissolved intetrabutylammonium flouride (120 mL of a 1.0M solution intetrahydrofuran, 0.124 mole). Ethyl bromoacetate (8.3 mL, 75 mmole) isadded. The reaction mixture is stirred at room temperature. The reactionmixture is concentrated, then partitioned between dichloromethane andwater. The aqueous layer is extracted with dichloromethane. The combinedorganic extracts are washed with water, brine, and dried over anhydrousmagnesium sulfate. The solvent is removed under vacuum to afford thetitle compound.

Example 43 Preparation of ethyl5-benzylsulfonylamino-1-methoxymethyl-3-uracilylacetate ##STR70##

A stirred solution of the compound of Example 42 (10.0 g, 35 mmole) inethanol (100 mL) is hydrogenated over 10% Pd/C (1.00 g) for 8 hoursunder balloon pressure. Celite is added, and the reaction mixture isfiltered through a pad of celite, using ethyl acetate to wash. Thesolvent is removed in vacuo to give crude ethyl5-amino-1-methoxymethyl-uracilylacetate.

A stirred solution of ethyl 5-amino-1-methoxymethyl-uracilylacetate (8.0g, 31 mmole) and 2,4,6-collidine (13.7 mL, 62 mmole) in tetrahydrofuran(50 mL) is cooled in an ice bath. A solution of benzylsulfonyl chloride(5.93 g, 31 mmole) in tetrahydrofuran (50 mL) is added over a 30-minuteperiod. After addition is complete, the solution is stirred for 1 hourat 0° C., then 3 hours at room temperature. The reaction mixture isdiluted with ethyl acetate, washed with 1.0N HCl (until aqueous layer ispH 1), water, saturated sodium bicarbonate, and brine. The organic layeris dried over anhydrous magnesium sulfate, and the solvent is removed.The title compound is isolated.

Example 44 Preparation of5-benzylsulfonylamino-1-methoxymethyl-3-uracilylacetic acid ##STR71##

To a cooled (0° C.) suspension of the compound of Example 43 (10.0 g, 24mmole) in methanol (50 mL) is added 1.0N NaOH (49 mL) over a period of10 minutes. After the addition is complete, the solution is allowed towarm to room temperature over a period of 1.5 hours. The solvent isreduced under vacuum, residue diluted with water, and washed with ethylacetate twice. The aqueous layer is acidified with 2.0N HCl to pH 1,extracted with ethyl acetate three times. The combined organic extractsare washed with water, then brine (twice). The solvent is removed togive the title compound.

Example 45 Preparation of5-benzylsulfonylamino-1-methoxymethyl-3-uracilylacetyl-N^(g)-nitro-L-argininal ethyl cyclol ##STR72##

To a stirred suspension of the compound of Example 44 (10.0 g, 26mmole), N^(g) -nitro-L-argininal ethyl cyclol, hydrochloride salt (8.38g, 31 mmole), and N-hydroxybenzotriazole (4.0 g, 26 mmole) inacetonitrile (200 mL) cooled to 0° C. is added1-ethyl-3-(3-dimethylamino-propyl)carbodiimide hydrochloride salt (6.0g, 31 mmole). N-methylmorpholine (8.6mL, 78 mmole) is added dropwise.After the addition is complete, the reaction is stirred at roomtemperature for 3 hours. The solvent is reduced under vacuum, and theresulting residue is dissolved in dichloromethane, washed with 2.0N HCl(to pH 1), water, saturated sodium bicarbonate and brine. The extract isdried over anhydrous magnesium sulfate, and the solvent is removed undervacuum. The crude product is chromatographed through silica gel to yieldthe title compound.

Example 46 Preparation of5-benzylsulfonylamino-1-methoxymethyl-3-uracilylacetyl-L-argininal ethylcyclol, acetate salt ##STR73##

The compound of Example 45 (1.00 g) in ethanol/acetic acid/water (4:1:1,10 mL) is hydrogenated over 10% palladium on carbon (0.30 g) for 4 hoursat 20 psi. Celite is added, and the solution is filtered through a 0.2micron filter, washing the solid with ethanol/acetic acid/water (4:1:1,10 mL). To the filtrate is added 10% palladium on carbon (0.30 g), andthe solution is hydrogenated at 20 to 25 psi until there is no startingmaterial as observed by analytical HPLC. Celite is added, and thesolution was filtered through a 0.2 micron filter, washing the solidwith water. The solvent is reduced to a volume of 80 mL under reducedpressure, then washed with ethyl acetate. The solvent from the aqueouslayer is reduced to remove the volatiles, then the sample is lyophilizedto yield the title compound.

Example 47 Preparation of5-benzylsulfonylamino-3-uracilylacetyl-L-argininal, trifluoroacetatesalt ##STR74##

The compound of Example 46 (1.00 g) is dissolved in 3.0N hydrochloricacid (20 mL). After 3 hours, the reaction mixture is quenched withaqueous sodium acetate (to pH 3.5), then filtered through a 2 micronfilter. The filtrate is purified by preparative HPLC (5×25 cm Vydac C-18column, 0 to 20% acetonitrile/water containing 0.1% trifluoroaceticacid). The clean fractions, as analyzed by analytical HPLC, are combinedto give the title compound.

Example 48 Preparation of 5-nitro-1-methyl-uracil ##STR75##

5-nitrouracil (10.00 g, 64 mmole), 1,1,1-3,3,3-hexamethyldisilazane (40mL, 190 mmole) and chlorotrimethylsilane (4.0 mL, 32 mmole) are heatedto reflux for 24 hours. The solution is concentrated under reducedpressure to afford 5-nitrouracil bis-(trimethylsilyl) ether.5-nitrouracil bis(trimethylsilyl) ether (10.0 g, 24 mmole),dimethylformamide (50 mL) and iodomethane (3.0 mL, 49 mmole) are heatedin an 80° C. oil bath for 24 hours. Ice water is added, and the mixtureis stirred for 30 minutes, then extracted with dichloromethane (3X). Thecombined organic layers are dried over magnesium sulfate, filtered, andconcentrated under reduced pressure to give the title compound. Analternative method of preparing the title compound is described inExample 108.

Example 49 Preparation of ethyl 5-nitro-1-methyl-3-uracilylacetate##STR76##

Using similar procedures to that described above in Example 42, thetitle compound is prepared from the compound of Example 48.

Example 50 Preparation of Ethyl5-benzylsulfonylamino-1-methyl-3-uracilylacetate ##STR77##

Using similar procedures to that described above in Example 43, thetitle compound is prepared from the compound of Example 49.

Example 51 Preparation of5-benzylsulfonylamino-1-methyl-3-uracilylacetic acid ##STR78##

Using similar procedures to that described above in Example 44, thetitle compound is prepared from the compound of Example 50. An alternatemethod of preparing the title compound is described in Example 111.

Example 52 Preparation of5-benzylsulfonylamino-1-methyl-3-uracilylacetyl-N^(g) -nitro-L-argininalethyl cyclol ##STR79##

Using similar procedures to that described above in Example 45, thetitle compound is prepared from the compound of Example 51.

Example 53 Preparation of5-benzylsulfonylamino-1-methyl-3-uracilylacetyl-L-argininal ethylcyclol, acetate salt ##STR80##

Using similar procedures to that described above in Example 46, thetitle compound is prepared from the compound of Example 52.

Example 54 Preparation of5-benzylsulfonylamino-1-methyl-3-uracilylacetyl-L-argininal,trifluoroacetate salt ##STR81##

Using similar procedures to that described above in Example 47, thetitle compound is prepared from the compound of Example 53. Analternative method of preparing the title compound is described inExample 113 (Compound E).

Example 55 Preparation ofalpha-N-benzyloxycarbonyl-omega,omega'-di-N-t-butoxycarbonyl-L-argininelactam ##STR82##

Alpha-N-t-benzyloxycarbonyl-omega,omega'-di-N-t-butoxycarbonylarginine(2.10 g. 4.1 mmole)) was dissolved in acetonitrile (25 mL).Hydroxybenzotriazole (0.63 g, 4.1 mmole) and1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride salt (0.79g, 4.1 mmole) were added in succession. After the reaction was stirredfor 1 hour, the solvent was reduced. The residue was dissolved in ethylacetate (50 mL), washed with water, saturated sodium bicarbonate, andbrine. The solution was dried over anhydrous magnesium sulfate,filtered, and the solvent was removed in vacuo to give 1.90 g (94%yield) of the title compound. R_(f) =0.37 (10% ethylacetate/dichloromethane).

Example 56 Preparation ofalpha-N-benzyloxycarbonyl-omega,omega'-di-N-t-butoxycarbonyl-L-argininal##STR83##

A solution of the compound of Example 55 (33.69 g, 69 mmole) intetrahydrofuran (350 mL) was cooled to -70° C. Lithium aluminum hydride(69 mL of a 1.0M solution in tetrahydrofuran) was added dropwise whilemaintaining the temperature below -60° C. After the reaction mixture wasstirred at -60 to -65° C. for 30 minutes, it was cooled to -70° C. 2.5 Mpotassium bisulfate (92 mL) was added dropwise to quench the excesslithium aluminum hydride. The solution was allowed to warm to 0° C., andthe mixture was filtered through celite, washing with ethyl acetate. Thefiltrate was washed with cold 1.0N HCl (75 mL), ice water (50 mL), coldsaturated sodium bicarbonate (50 mL), then cold brine (50 mL). Theextract was diluted with dichloromethane (200 mL), then dried overanhydrous magnesium sulfate. The solvent was removed in vacuo to afford28.23 g. (83%) of the title compound. R_(f) =0.37 (5% isopropanol indichloromethane).

Example 57 Preparation of omega,omega'-di-N-t-butoxycarbonyl-L-argininal diethyl acetal, HCl salt##STR84##

The compound of Example 56 (300 mg, 0.61 mmole) was dissolved in ethanol(3.0 mL) and concentrated HCl was added (51 microliters). After stirringovernight at room temperature, 10% Pd/C (30 mg) was added. The mixturewas hydrogenated for 4 hours. TLC indicated that the reaction wascomplete. Celite was added, and the reaction mixture was filtered. Thesolution was diluted with water to a volume of 50 mL. The title compound(190 mg, 73% yield) was freeze dried to a yellow solid. R_(f) =0.26 (10%methanol/dichloromethane).

Example 58 Preparation of t-butyl(3-nitro-2-oxo-1,2-dihydropyridyl)acetate ##STR85##

Sodium hydride (1.57 g of a 60% dispersion in mineral oil, 0.039 mole)was washed with hexanes three times (10 mL each) and suspended indimethylformamide (25 mL). The stirred suspension was cooled in an icebath, then 3-nitro-2-hydroxypyridine (5.00 g, 0.036 mole) was added insmall portions over a 25-minute period. After the addition was complete,the reaction was stirred at 0° C. for 10 minutes, then room temperaturefor 30 minutes. The reaction mixture was recooled in an ice bath.t-Butyl bromoacetate (5.25 mL, 0.036 mole) was added. The reaction wasstirred at 0° C. for 1 hour, then 1.5 hours at room temperature. Thereaction mixture was diluted with ethyl acetate (80 mL), and ice (80 g)was added. The aqueous layer was extracted with ethyl acetate (3×200mL). The combined organic extracts were washed with water (4×100 mL),brine (100 mL), and dried over anhydrous sodium sulfate. The solvent wasremoved, and the resulting residue was chromatographed through silicagel using 10% ethyl acetate/dichloromethane as eluent. The purefractions were combined, and the solvent was removed under vacuum toafford 6.77 g (75% yield) of the title compound as yellow solid. R_(f)=0.30 (silica gel, 20% ethyl acetate in dichloromethane).

Example 59 Preparation of t-butyl(3-amino-2-oxo-1,2-dihydropyridyl)acetate ##STR86##

A stirred solution of the compound of Example 58 (2.00 g, 7.9 mmole) inethanol (50 mL) was hydrogenated over 10% Pd/C (0.23 g) for 3 hoursunder balloon pressure. Celite was added, and the reaction mixture wasfiltered through a pad of Celite, using methanol/ethyl acetate to wash.The solvent was removed under vacuum to afford the title compound (1.90g) in quantitative yield. R_(f) =0.56 (silica gel, 10% methanol indichloromethane).

Example 60 Preparation of ethyl3-[(allyloxycarbonyl)amino]-2-oxo-1,2-dihydropyridylacetate ##STR87##

The compound of example 59 (1.7 g, 7.85 mmole) is dissolved in 50%aqueous dioxane (30 mL) and cooled to 0° C. Sodium bicarbonate (2.0 g,24 mmole) is added in one portion. After stirring 5 minutes, allylchloroformate (1.67 g, 16 mmole) in dioxane (4 mL) is added dropwiseover a 5-minute period. After stirring for 30 minutes, the solvent isreduced to a volume of 10 mL, and extracted with dichloromethane (50mL). The organic layer is washed with brine, the dried over anhydrousmagnesium sulfate. The solvent is removed and the title compoundisolated.

Example 61 Preparation of3-[(allyloxycarbonyl)amino]-2-oxo-1,2-dihydropyridylacetic acid##STR88##

The compound of example 60 (1.00 g) is treated with 50% trifluoroaceticacid in dichloromethane (10 mL) for 1 hour at 0° C., and 3 hours at roomtemperature. The solution is diluted with toluene (50 mL) and thesolvent is removed in vacuo to afford the title compound.

Example 62 Preparation of3-[(allyloxycarbonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-omega,omega'-di-N-t-butoxycarbonyl-L-argininal diethyl acetal ##STR89##

To a stirred suspension of the compound of Example 57 (500 mg, 1.98mmole), the compound of example 61 (1.12 g, 2.37 mmole), andN-hydroxybenzotriazole (300 mg, 1.98 mmole) cooled to 0° C., is added1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride salt (457mg, 2.38 mmole). N-methylmorpholine (0.65 mL, 5.9 mmole) is addeddropwise. After the addition is complete, the reaction is stirred atroom temperature overnight. The solvent is reduced under vacuum, and theresulting residue is dissolved in ethyl acetate, washed with 1.0N HCl(to pH 1), water, saturated sodium bicarbonate and brine. The extract isdried over anhydrous magnesium sulfate, and the solvent is removed undervacuum to yield the title compound.

Example 63 Preparation of3-[(allyloxycarbonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,trifluoroacetate salt ##STR90##

The compound of example 62 (1.0 g) is suspended in 50% aqueousacetonitrile (20 ml) and cooled in an ice bath. Hexafluorophosphoricacid (60% by weight, 10 mL) is added slowly, and the cooling bath isremoved. After 30 minutes, the reaction mixture is recooled in an icebath, and quenched with aqueous sodium acetate (2.5M solution) to pH 4,then filtered through a 2 micron filter. The filtrate is purified usingpreparative HPLC. The fractions are analyzed for purity by analyticalHPLC (0.1% trifluoroacetic acid/10-40% aqueous acetonitrile), combined,and the acetonitrile is removed under reduced pressure. The remainingwater is lyophilized. The title compound is recovered.

Example 64 Preparation of N-(t-butoxycarbonyl)-3-(3-pyridyl)-L-alaninemethyl ester ##STR91##

To a solution of N-(t-butoxycarbonyl)-3-(3-pyridyl)alanine (5.0 g, 18.8mmole) in methanol (100 mL) was added thionyl chloride (2M solution indichloromethane, 66 mL, 132 mmole). The resulting solution was stirredovernight at ambient temperature. The methanol was removed under reducedpressure to a minimum volume and ethyl acetate (100 mL) was added. Theresulting white precipitate was collected in a fritted funnel. To asolution of the collected precipitate in a mixture oftetrahydrofuran/water (40 mL each) was added di-tert-butyl dicarbonate(4.8 g, 21.99 mmole) and sodium carbonate (1.95 g, 18.4 mmole). Afterstirring for 12 hours at ambient temperature, the reaction mixture wasdiluted with ethyl acetate (40 mL) and washed with a solution ofsaturated sodium bicarbonate (25 mL). The organic layer was dried overanhydrous sodium sulfate and concentrated under vacuum to give crudeproduct. This product was subjected to flash column chromatography onsilica gel (230-400 mesh) using a 8×52 cm column and eluting with a10:90 mixture of ethyl acetate/hexane followed by a 60:40 mixture ofethyl acetate/hexane. 4 g (74%) of the title compound was obtained as anoil. Thin-layer chromatography gave a R_(f) =0.68 (silica gel; ethylacetate).

Example 65 Preparation of N-(t-butoxycarbonyl)-3-(3-piperidyl)-L-alaninemethyl ester, acetate salt ##STR92##

A solution of the compound of Example 64 (5 g, 17.8 mmole) in ethanol(24 mL), acetic acid (6 mL) and water (6 mL) was hydrogenated overplatinum oxide (500 mg) at 45 psi for three hours. The catalyst wasfiltered off and the filtrate concentrated under vacuum to an oilyresidue (6.89 g) which was used in the next step (Example 6) withoutfurther purification. Thin-layer chromatography yielded two spotscorresponding to two diastereomers with R_(f) values of 0.16 and 0.26,respectively (silica gel; 4:1:1 n-butanol/acetic acid/water).

Example 66 Preparation of N-(t-butoxycarbonyl)-3-[3-piperidyl-(N-guanidino (bis-benzyloxycarbonyl))]-L-alanine methylester ##STR93##

To a solution of the compound of Example 65 (6.89 g, 19.9 mmole) intetrahydrofuran (80 mL) was added S-methylisothioureabis-benzyloxycarbonyl (7.13 g, 19.9 mmole) followed byN-methylmorpholine (4.37 mL), and the reaction mixture was stirred atambient temperature for 18 hours. The reaction mixture then wasconcentrated under vacuum and the resulting residue was dissolved inethyl acetate (100 mL) and washed with 1N sodium bisulfate and saturatedsodium chloride (50 mL each). After drying over anhydrous sodiumsulfate, the solvents were removed under vacuum; the crude titlecompound was subjected to flash column chromatography on silica gel(230-400 mesh) using a 8×52 cm column and eluting with 1:9 ethylacetate/hexanes (two column volumes) followed by 1:1 ethylacetate/hexanes. 2.75 g the title compound was obtained as a mixture oftwo diastereomers. Thin-layer chromatography gave two spots with R_(f)values of 0.57 and 0.62, respectively (silica gel; 1:1 ethylacetate/hexanes).

Example 67 Preparation ofN-(t-butoxycarbonyl)-3-[3-piperidyl-(N-guanidino(bis-benzyloxycarbonyl))]-L-alaninol ##STR94##

To a stirred solution of the compound of Example 66 (2.23 g, 3.7 mmole)in absolute ethanol (8 mL) and anhydrous tetrahydrofuran (4 mL) wasadded calcium chloride (844 mg, 7.6 mmole) and sodium borohydride (575mg, 15.2 mmole). After stirring 12 hours at ambient temperature, thereaction mixture was concentrated under vacuum and the resulting residuewas partitioned between ethyl acetate and 1N sodium bisulfate (10 mLeach). The two layers were separated; organic layer was washed twicemore with 1N sodium bisulfate, dried over anhydrous sodium sulfate andconcentrated under vacuum gave a residue. Flash column chromatography ofthe residue on silica gel (230-400 mesh) using a 5.5×45 cm column andeluting with ethyl acetate gave 1.3 g of the title compound as a whitefoam. Thin layer chromatography yielded two spots corresponding to twodiastereomers with R_(f) values of 0.18 and 0.27, respectively (silicagel; 1:1 ethyl acetate/hexanes).

Example 68 Preparation of3-[3-piperidyl-(N-guanidino(bis-benzyloxycarbonyl))]-L-alaninol,hydrochloride salt ##STR95##

The compound of Example 67 (290 mg, 0.57 mmole) was treated with 2.5 Nanhydrous hydrochloric acid in ethyl acetate (2.0 mL) at ambienttemperature for one hour. The solvent was removed under vacuum to give asticky-white solid (260 mg) . This solid was used in the next step(Example 20) without further purification. ¹ H NMR spectrum taken in CD₃OD showed no t-butoxycarbonyl protons at 1.4 ppm.

Example 69 Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-3-[3-piperidyl-(N-quanidino(bis-benzyloxycarbonyl))]-alaninol##STR96##

To a suspension of the compound of Example 68 (266 mg, 0.45 mmole) inacetonitrile (7 mL) is added successively the compound of Example7[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetic acid (145mg, 0.41 mmole), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimidehydrochloride salt (86 mg, 0.45 mmole), 1-hydroxybenzotriazole hydrate(72 mg, 0.47 mmole) and diisopropylethylamine (2.44 mmole, 417microliters). The solution is stirred at ambient temperature for twelvehours. The solvent is removed under reduced pressure and the remainingresidue is dissolved in ethyl acetate (15 mL) and washed two times eachwith 10 mL portions of 1N sodium bisulfate, saturated sodium bicarbonateand saturated sodium chloride solution. The organic layer is dried oversodium sulfate and concentrated to crude product. The title compound isisolated.

Example 70 Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-3-[3-piperidyl-(N-guanidino)]alaninol,acetate salt ##STR97##

The compound of Example 69 (123 mg, 0.16 mmole) is subjected tocatalytic hydrogenation in methanol (8 mL), and acetic acid (2 mL) andwater (2 mL) in the presence of palladium on carbon (20 mg) at 40 psifor 4 hours. The title compound is obtained.

Example 71 Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-3-[3-piperidyl-(N-guanidino)]alaninal##STR98##

To a chilled solution of the compound of Example 70 (107 mg, 0.19 mmole)in dimethylsulfoxide and toluene (2 mL each) is added dichloroaceticacid (78 microliter, 0.94 mmole) followed by1-ethyl-3-(3-dimethylamino-propyl)carbodiimide hydrochloride salt (0.36g, 1.9 mmole) at one minute later. The reaction is stirred for 5 minutesat 0° C. and 85 minutes at ambient temperature, and quenched with 30 mLwater. The water layer is extracted twice with diethyl ether (15 mLportions), diluted to 60 mL with water and is purified by high pressureliquid chromatography using a reverse phase column containing a C-18resin comprised of 10 micron-size gel particles with a 300 angstrom poresize. The column is eluted with a water/acetonitrile (containing 0.1%trifluoroacetic acid) gradient where the gradient is run from 10% to 30%acetonitrile. Each diasteromer of the title compound is isolated.

Example 72 Preparation semicarbazid-4-yl diphenylmethane,trifluoroacetate salt ##STR99## Step 1

A solution of carbonyldiimidazole (16.2 g, 0.10 mole) in 225 mL ofdimethylformamide was prepared at room temperature and allowed to stirunder nitrogen. A solution of t-butyl carbazate (13.2 g, 0.100 moles) in225 mL dimethylformamide was then added dropwise over a 30 minuteperiod. Next, diphenylmethylamine (18.3 g, 0.10 moles) was added over a30 minute period. The reaction was allowed to stir at room temperatureunder nitrogen for one hour. Water (10 mL) was added and this mixturewas concentrated to about 150 mL under vacuum. This solution was pouredinto 500 mL water and extracted with 400 mL of ethyl acetate. The ethylacetate phase was extracted two times each with 75 mL 1N HCl, water,saturated sodium bicarbonate and brine, and then was dried withanhydrous magnesium sulfate. The mixture was filtered and the solutionwas concentrated to give 29.5 g (85% yield) of1-t-butoxycarbonyl-semicarbazid-4-yl diphenylmethane as a white foam.This material may be purified by recrystallization from ethylacetate/hexane, but was pure enough to use directly in step 2: mp142-143° C. ¹ H NMR (CDCl₃) delta 1.45 (s, 9H), 6.10 (dd, 2H), 6.42 (s,1H), 6.67 (bs, 1H), 7.21-7.31 (m, 10H). Analysis Calculated for C₁₉ H₂₃N₃ O₃ : C, 66.84; H, 6.79; N, 12.31. Found: C, 66.46; H, 6.75; N; 12.90.

Step 2

A solution of 3.43 g (10 mmole) of 1-t-butoxycarbonyl-semicarbazid-4-yldiphenylmethane in 12.5 mL of dichloromethane was treated with 12.5 mLof trifluoroacetic acid at 0° C. The reaction mixture was allowed tostir for 30 minutes at this temperature. The reaction mixture was thenadded dropwise to 75 mL of diethyl ether to give a precipitate. Theresulting precipitate was filtered off and washed with diethyl ether togive 2.7 g (80% yield) of the title compound; mp 182-184° C.

Example 73 Preparation of 3-thioamidobenzyl-N-acetylaminomalonic aciddiethyl ester ##STR100##

To a stirred solution of alpha-bromo-meta-tolunitrile (45.0 g, 0.24mole), diethyl acetamidomalonate (48.0 g, 0.22 mole) and potassiumiodide ((3.0 g, 0.018 mole) in dioxane (500 mL) was added 2.5M sodiumethoxide in ethanol (100 mL) dropwise under an argon atmosphere. Afterthe addition was complete, the solution was refluxed for 6 hours. Thereaction mixture was allowed to stand overnight at room temperature,then diluted with brine (250 mL) and water (250 mL), and extracted withethyl acetate four times (1.0 L total). The combined extracts werewashed with water (100 mL), 10% citric acid (100 mL), water (100 mL) andbrine (2×50 mL), then dried over anhydrous magnesium sulfate andfiltered; the solvent was removed under vacuum. The crude residue wasrecrystallized from ethyl acetate and diethyl ether in two crops toyield 43.51 g (60%) of the 3-cyanobenzyl-N-acetylaminomalonic aciddiethyl ester as yellow crystals.

H₂ S(g) was bubbled into a rapidly stirring solution of3-cyanobenzyl-N-acetylaminomalonic acid diethyl ester (44.3 g, 0.13mmole) in pyridine (300 mL) and triethylamine (100 mL) for 40 minutes.The reaction mixture was stirred at room temperature for 16 hours, thenpoured into 3.0 L of water. A yellow precipitate formed immediately. Thesolution was allowed to stand at 4° C. for 4 hours, then was filtered.The crude title compound was recrystallized from ethyl acetate andhexanes to yield 48.1 g (98%) of the title compound as yellow crystals.m.p. 183-186° C. ¹ H NMR (CDCl₃): delta 1.31 (t, J=7.1 Hz, 6H), 2.06 (s,3H), 3.70 (s, 2H), 4.29 (q, J=7.1 Hz, 4H), 4.80-4.87 (m, 1H), 6.60 (s,1H), 7.10-7.20 (m, 1H), 7.27-7.35 (m, 2H), 7.60-7.70 (m, 2H). AnalysisCalculated for C₁₇ H₂₂ N₂ O₅ S: C, 55.72; H, 6.05; N, 7.64. Found: C,55.55; H, 5.96; N, 7.76.

Example 74 Preparation of 3-amidino-D,L-phenylalanine, dihydrochloridesalt ##STR101##

The compound of Example 73 (48.1 g, 0.13 mmole) was dissolved in acetone(800 mL). Iodomethane (18.3 mL, 0.19 mole, 1.5 equivalents) was added,and the solution was refluxed for 30 minutes. The solution was cooled toroom temperature, and the intermediate thioimidate was filtered, driedand dissolved in methanol (500 mL). Ammonium acetate (14.8 g, 0.19 mole,2 equivalents) was added. The reaction mixture was refluxed for 1 hour,then cooled to room temperature, and poured into ether (1.2 L). Thesolution was allowed to stand at 4° C. for 72 hours. The crude3-amidinobenzyl-N-acetylaminomalonic acid diethyl ester was filtered,washed with ether, air dried, and then refluxed in concentrated HCl (250mL) for 3 hours. The reaction mixture was concentrated under vacuum,diluted with water (0.5 L), and concentrated under vacuum again. Thesesteps were repeated. The crude title compound was purified bycation-exchange (Sephadex SP-C25) using a gradient of 0-0.1N HCl aseluent to yield 10.8 g (30%) of the title compound as an off-whitesolid. ¹ H NMR (D₂ O): delta 3.14-3.29 (2H, m), 4.17 (dd, J=7.4, 6.2 Hz,1H), 7.42-7.69 (4H, m). Analysis Calculated for C₁₀ H₁₃ N₃ O₂.2HCl.1.9H₂O: C, 38.20; H, 6.03; N, 13.36. Found: C, 38.51; H, 5.64; N, 12.89.

Example 75

Preparation ofN-alpha-Boc-N-omega-4-methoxy-2,3,6-trimethylbenzenesulfonyl-3-amidino-D,L-phenylalanine##STR102##

The compound of Example 74 (3-amidino-D,L-phenylalanine) (4.00 g, 13mmole) was dissolved in 50% aqueous dioxane (20 mL). Sodium bicarbonate(3.38 g, 40 mmole) was added, followed by di-t-butyl dicarbonate (2.93g, 13 mmole) in dioxane (4 mL). The reaction mixture was stirred for 18hours at room temperature. The solution was cooled in an ice bath, and4.0 N sodium hydroxide was added dropwise until the solution was pH 12.4-methoxy-2,3,6-trimethylbenzenesulfonyl chloride (8.01 g, 32 mmole) indioxane (10 mL) was added dropwise. 4.0 N sodium hydroxide was added asneeded to keep the pH at 12. The ice bath was removed. After 1 hour, 1.0N HCl was added to bring the solution to pH 7 to 8. The solution wasdiluted with an additional 50 mL of water and then was washed with ethylacetate two times (20 mL each). The aqueous layer was acidified to pH1.0 with 1.0 N HCl and extracted with ethyl acetate three times (100 mLtotal). The combined organic layers were washed with water (20 mL) andbrine twice (10 mL each). The organic layer was dried over anhydrousmagnesium sulfate and the solvent was removed under vacuum. The residuewas dissolved in a minimum amount of dichloromethane, then addeddropwise to ether (25 mL). Solid impurities were removed by filteringand the solvent removed from the filtrate under vacuum to give 4.90 g(68% crude yield) of the title compound as an off-white foam. A 30 mgsample of the title compound was further purified by preparativethin-layer chromatograph developing with 1% acetic acid/5%isopropanol/dichloromethane to give 9 mg of the title compound in apurer form. Rf=0.16 (1% acetic acid/5% isopropanol/dichloromethane). ¹ HNMR (CD₃ OD): delta 1.32 (s, 9H), 2.14 (s, 3, H), 2.63 (s, 3H), 2.71 (s,3H), 2.93 (dd, J=13.7, 9.3 Hz, 1H), 3.22 (dd, J=13.7, 4.3 Hz, 1H), 3.85(s, 3H), 4.34-4.37 (m, 1H), 6.72 (s, 1H), 7.35-7.47 (2H, m), 7.69-7.75(m, 2H).

Example 76

Preparation ofN-alpha-Boc-N-omega-4-methoxy-2,3,6-trimethylbenzenesulfonyl-3-amidino-D,L-phenylalanine-N-methyl-O-methyl-carboxamide##STR103##

To a stirred solution of compound of Example 75 (1.00 g, 1.92 mmole),O,N-dimethyl hydroxylamine hydrochloride (375 mg, 3.85 mmole),hydroxybenzotriazole hydrate (294 mg, 1.92 mmole) and 4-methylmorpholine(1.06 mL, 9.62 mmole) in tetrahydrofuran (4 mL), cooled in an ice bath,was added 1-ethyl-3-(3-dimethylamino-propyl)carbodiimide hydrochloridesalt (406 mg, 2.12 mmole). The ice bath was removed, and the reactionmixture was stirred for 2 hours at room temperature. The reactionmixture was diluted with ethyl acetate (75 mL), washed with water, 10%citric acid, water, saturated sodium bicarbonate, and brine. The organiclayer was dried over anhydrous magnesium sulfate and the solvent wasremoved under vacuum. 750 mg (69%) of the title compound was isolated. ¹H NMR (CDCl₃): delta 1.33 (s, 9H), 2.14 (s, 3H), 2.66 (s, 3H), 2.75 (s,3H), 2.80-2.88 (m, 1H), 3.06-3.20 (m, 4H), 3.70 (s, 3H), 3.84 (s, 3H),4.98-5.06 (m, 1H), 5.21 (d, J=8.7 Hz, 1H), 6.48 (bs, 1H), 6.58 (s, 1H),7.30-7.34 (m, 2H) 7.60-7.68 (m, 2H), 8.11 (bs, 1H). Analysis Calculatedfor C₂₇ H₃₈ N₄ O₇ S.0.5H₂ O: C, 56.73; H, 6.88; N, 9.80. Found: C,56.97; H, 6.66; N, 9.43.

Example 77

Preparation ofN-alpha-Boc-N-omega-4-methoxy-2,3,6-trimethylbenzenesulfonyl-D,L-3-amidinophenylalaninal##STR104##

To a stirred solution of LiAlH₄ (2.00 mL of a 1.0 M solution intetrahydrofuran, 1.24 mmole) in tetrahydrofuran (8 mL), cooled in a dryice/acetone bath, the compound of Example 76 (0.75 g, 1.9 mmole intetrahydrofuran (5 mL)) was added dropwise. The cooling bath was removedand the reaction mixture was allowed to warm to 5° C. The reactionmixture was re-cooled in the dry ice acetone bath and quenched with 3.0mL of a 1:2.7 wt./wt. solution of potassium bisulfate in water. Thereaction mixture was allowed to warm to room temperature, stirred for 3hours, filtered and concentrated under vacuum. The residue was dissolvedin ethyl acetate (20 mL), and washed with 10% citric acid (2 mL), water(2 mL), saturated sodium bicarbonate (2 mL) and brine (2 mL). Theorganic layer was dried over anydrous magnesium sulfate and the solventwas removed under vacuum to yield 580 mg (86%) of the title compound. ¹H NMR (CDCl₃): delta 1.31 (s, 9H), 2.07 (s, 3H), 2.57 (s, 3H), 2.67 (s,3H),2.90-3.17 (2H, m), 3.77 (s, 3H), 4.33-4.40 (1H, m), 5.02-5.08 (1H,m), 6.48 (1H, s), 7.23-7.31 (2H, m), 7.50-7.62 (2H, m), 7.94, (1H, bs),8.05 (1H, bs), 9.55 (1H, s). Analysis Calculated for C₂₅ H₃₃ N₃ O₆S.0.5H₂ O: C, 58.58; H, 6.69; N,8.20. Found: C, 58.57; H, 6.72; N, 7.98.

Example 78

Preparation ofN-alpha-Boc-N-omega-4-methoxy-2,3,6-trimethylbenzenesulfonyl-D,L-3-amidinophenylalaninal-semicarbazonyl-4-N-diphenylmethane##STR105##

The compound of Example 77 (0.58 g, 1.9 mmole), the compound of Example72 (410 mg, 1.15 mmole) and sodium acetate trihydrate (188 mg, 1.38mmole) were refluxed in 75% aqueous ethanol (10 mL) for 1 hour. Afterthe reaction mixture was cooled to room temperature, it was diluted withethyl acetate (50 mL), washed with l.ON HCl (5 mL), water (5 mL),saturated sodium bicarbonate (5 mL) and brine (2×5 mL), and dried overanhydrous magnesium sulfate. The solvent was removed under vacuum toyield 750 mg (89% yield) of the title compound as an off-white foam.Analysis calculated for C₃₉ H₄₆ N₆ O₆ S.1.0H₂ O: C, 62.88; H, 6.49; N,11.28. Found: C, 63.14; H, 6.35 N, 11.10.

Example 79

Preparation of N-omega-4-methoxy-2,3,6-trimethylbenzenesulfonyl-D,L-3-amidinophenylalaninal-semicarbazonyl-4-N-diphenylmethane,trifluoroacetate salt ##STR106##

The compound of Example 78 (750 mg, 1.9 mmole) was treated with 50%trifluoroacetic acid/dichloromethane (3 mL) for 30 minutes at roomtemperature. The reaction mixture was added dropwise to ether (50 mL).The solution was allowed to stand at 4° C. for 18 hours. The product wasfiltered, and dried under vacuum to yield 600 mg (79% yield) of thetitle compound as an off-white solid. Analysis calculated for C₃₉ H₄₆ N₆O₆ S.1.3CF₃ CO₂ H: C, 56.72; H, 5.11; N, 10.84. Found: C, 56.34; H,5.47; N, 11.49.

Example 80

Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-D,L-N-omega-4-methoxy-2,3,6-trimethylbenzenesulfonyl-D,L-3-amidinophenylalaninal-semicarbazonyl-4-N-diphenylmethane ##STR107##

1-Ethyl-3-(3-dimethylamino-propyl)carbodiimide hydrochloride salt (94mg, 0.94 mmole) is added in one portion to a stirred solution of thecompound of Example 7 (303 mg, 0.49 mmole), hydroxybenzotriazole (75 mg,0.49 mmole), and 4-methylmorpholine (0.24 mL, 2.2 mmole) indimethylformamide (5 mL) with cooling in an ice bath. After 30 minutes,the compound of Example 79 (363 mg, 0.49 mmole) is added. After anadditional 2 hours, the reaction mixture is diluted with water (25 mL)and brine (25 mL). The product is filtered and dissolved into ethylacetate (25 mL). The solution is washed with 10% citric acid, water,saturated sodium bicarbonate and brine, and is dried over anhydrousmagnesium sulfate. The solvent is removed under vacuum. The resultingresidue is chromatographed by flash chromatography on silica gel to givethe title compound.

Example 81

Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-D,L-3-amidinophenylalaninal semicarbazone ##STR108##

The compound of Example 80 (102 mg, 0.11 mmole) is treated withhydrofluoric acid/anisole (9:1) for 30 minutes at -20° C. and 0° C. for30 minutes. After removal of the hydrofluoric acid, the resultingresidue is dissolved in 20% aqueous acetic acid and washed with diethylether. The aqueous layer is lyophilized to a powder, then is purified bypreparative HPLC (C-18, eluting with 10 to 40% acetonitrile-watergradient containing 0.1% trifluoroacetic acid) to give the titlecompound.

Example 82

Preparation of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-D,L-3-amidinophenylalaninal ##STR109##

The compound of Example 81 (16.6 mg, 30 micromole) is dissolved inmethanol (1 mL) and 1% aqueous trifluoroacetic acid (5 mL), thenformalin (0.23 mL) is added. After 40 minutes, the solution is filteredthrough a 2 micron filter, diluted to a volume of 15 mL with water, andthen is purified by preparative HPLC (C-18, eluting with 10 to 40%acetonitrile-water gradient containing 0.1% trifluoroacetic acid). Thefractions containing the title compound are pooled and lyophilized togive the title compound.

Example 83

Preparation ofethyl(3-[(N-t-butyloxycarbonyl)amino]-2-oxo-1,2-dihydropyridyl)acetate##STR110##

A stirred solution of the compound of Example 5 (44.5 g, 0.197 mole) inethanol (200 mL) was hydrogenated over 10% Pd/C (2.25 g) for 16 hoursunder balloon pressure. Celite was added, and the reaction mixture wasfiltered through a pad of celite in a 600 mL fritted funnel (5 cmdepth), using ethyl acetate to wash. The solvent was removed undervacuum, diluted with ethyl acetate (200 mL) and toluene (200 mL), andthe solvent was removed under vacuum to give crudeethyl(3-amino-2-oxo-1,2-dihydropyridyl)acetate (40.0 g, 0.204 mol) inquantitative yield.

A stirred solution of ethyl(3-amino-2-oxo-1,2-dihydropyridyl)acetate(2.00 g, 10 mmol) and sodium bicarbonate (1.69 g, 9.5 mmol) in 50%aqueous dioxane (20 ml) is cooled in an ice bath. A solution ofdi-t-butyldicarbonate (2.08 g, 20 mmol) in dioxane (10 ml) is added overa 5 minute period. After addition is complete, the solution is stirredfor 16 hours at room temperature. The reaction mixture is diluted withethyl acetate (100 mL), washed with 1.0N HCl (until aqueous layer is pH1), water, saturated sodium bicarbonate, and brine. The organic layer isdried over magnesium sulfate, and the solvent is removed. The titlecompound is isolated.

Example 84

Preparation ofethyl(3-[(N-t-butyloxycarbonyl-N-methyl)amino]-2-oxo-1,2-dihydropyridyl)acetate##STR111##

The compound of Example 83 (3.00 g, 10 mmol) and iodomethane (1.2 mL, 20mmol) are dissolved in tetrahydrofuran (30 mL), and the solution iscooled to 0° C. under a nitrogen atmosphere. Sodium hydride (0.44 g of a60% dispersion in mineral oil, 11 mmol) is added cautiously with gentlestirring. After the addition is complete, the reaction mixture isstirred at room temperature for 16 hours. Ethyl acetate (50 mL) isadded, followed by water, to destroy the excess sodium hydride. Theorganic layer is washed with water, 5% aqueous sodium thiosulfate (toremove the iodine), water and brine, dried over magnsium sulfate, andevaporated. The title compound is isolated.

Example 85

Preparation of(3-[(N-t-butyloxycarbonyl-N-methyl)amino]-2-oxo-1,2-dihydropyridyl)aceticacid ##STR112##

To a cooled (0° C.) suspension of the compound of Example 84 (3.2 g, 10mmole) in methanol (10 mL) is added 1.0N NaOH (12 ml) over a period of10 minutes. After the addition is complete, the solution is allowed towarm to room temperature over a period of 3 hours. The solvent isreduced under vacuum, the residue is diluted with water (25 mL), andwashed with ethyl acetate. The aqueous layer was acidified with 2.0N HClto pH 1, extracted with ethyl acetate three times. The combined organicextracts are washed with water, then brine (twice). The solvent isremoved, and the title compound is isolated.

Example 86

Preparation of[3-(N-t-butyloxycarbonyl-N-methyl)amino-2-oxo-1,2-dihydropyridyl]acetyl-N.sup.g-nitro-L-argininal ethyl cyclol ##STR113##

To a stirred suspension of the compound of Example 85 (2.2 g, 7.7mmole), N^(g) -nitro-L-argininal ethyl cyclol, hydrochloride salt (2.47g, 9.2 mmol), and N-hydroxybenzotriazole (1.17 g, 7.7 mmole) cooled to0° C. is added EDC (1.77 g, 9.2 mmole). N-methylmorpholine (2.5 mL, 23mmole) is added dropwise. After the addition is complete, the reactionis stirred at room temperature for 3 hours. The solvent is reduced undervacuum, and the resulting residue is dissolved in dichloromethane,washed with 2.0N HCl (to pH1), water, saturated sodium bicarbonate andbrine. The extract is dried over magnesium sulfate, and the solvent isremoved under vacuum. The title compound is isolated.

Example 87

Preparation of[3-(N-t-butyloxycarbonyl-N-methyl)amino-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalethyl cyclol, acetate salt ##STR114##

The compound of Example 86 (5.5 g, 11 mmole) in ethanol/aceticacid/water (4:1:1, 60 mL) is hydrogenated over 10% palladium on carbon(1.80 g) for 4 hours at 20 psi. Celite is added, and the solution isfiltered through a 0.2 micron filter, washing the solid withethanol/acetic acid/water (4:1:1, 60 mL). To the filtrate is added 10%palladium on carbon (1.80 g), and the solution is hydrogenated at 20-25psi for 40 hours. Celite is added, and the solution is filtered througha 0.2 micron filter, washing the solid with water (200 mL). The solventis reduced to a volume of 200 mL under reduced pressure, then washedwith ethyl acetate (50 mL) . The solvent from the aqueous layer isreduced to remove the volatiles, then the sample is lyophilized to yieldthe title compound.

Example 88

Preparation of[3-(N-t-butyloxycarbonyl-N-methyl)amino-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,trifluoroacetate salt ##STR115##

The compound of Example 87 (4.7 g, 9.2 mmole) is suspended in 3.0N HCl(100 mL) is added. After stirring for 3 hours, the reaction mixture isquenched with 2.5 M aqueous sodium acetate to pH 3.5 to 4, then filteredthrough a 2 micron filter. The filtrate is purified by preparative HPLC(Waters PrepPak cartridge, Delta-Pak C18, 300 angstrom column, 0-40%acetonitrile/water containing 0.1% trifluoroacetic acid). The cleanfractions are combined to give the title compound.

Example 89

General Procedure for Reaction of Ethyl(3-amino-2-oxo-1,2-dihydropyridyl)acetate with sulfonyl or sulfamoylchlorides

To a stirred solution of ethyl (3-amino-2-oxo-1,2-dihydropyridyl)acetate (5.89 g, 30 mmole) in dry tetrahydrofuran (300 mL) is added the2,4,6-collidine (7.93 mL, 60 mmol) and the solution is cooled to 0° C.under nitrogen. The appropriate sulfonyl or sulfamoyl chloride listedbelow (33 mmol) dissolved in tetrahydrofuran (25-75 mL) is addeddropwise. After the addition is complete, the reaction mixture isstirred for 30 minutes to 1 hour at 0° C. and then at ambienttemperature for 0 to 72 hours. The reaction mixture is diluted withethyl acetate, washed successively with 1.0N HCl, water, saturatedsodium bicarbonate and brine, dried over magnesium sulfate, and thesolvent is removed in vacuo. The residue is chromatographed on silicagel using a gradient system of dichloromethane and 1-4% methanol indichloromethane to afford the product, judged pure by TLC (silica gel).Using this method and the starting materials listed below, intermediateshaving the formula given below are made:

    ______________________________________                                          #STR116##                                                                   R =             Starting Material                                             ______________________________________                                        phenyl          benzenesulfonyl chloride                                        1-naphthyl 1-naphthylsulfonyl chloride                                        2-naphthyl 2-naphthylsulfonyl chloride                                        2-carbomethoxyphenyl 2-carbomethoxybenzenesulfonyl                             chloride                                                                     2-trifluoromethylbenzyl 2-trifluoromethylbenzylsulfonyl                        chloride                                                                     2-cyclohexylamino cyclohexylsulfamoyl chloride                                2-trifluoromethylphenyl 2-trifluoromethylbenzenesulfonyl                       chloride                                                                     3-trifluoromethylphenyl 3-trifluoromethylbenzenesulfonyl                       chloride                                                                     4-trifluoromethylphenyl 4-trifluoromethylbenzenesulfonyl                       chloride                                                                     2-methylphenyl 2-methylbenzenesulfonyl chloride                               3-methylphenyl 3-methylbenzenesulfonyl chloride                               2-methyl-5-fluorophenyl 2-methyl-5-fluorobenzenesulfonyl                       chloride                                                                     2-methoxyphenyl 2-methoxybenzenesulfonyl                                       chloride                                                                     3-methoxyphenyl 3-methoxybenzenesulfonyl                                       chloride                                                                     2-methoxy-5-chlorophenyl 2-methoxy-5-chlorobenzene-                            sulfonyl chloride                                                            2-nitrophenyl 2-nitrobenzenesulfonyl chloride                                 2-trifluoromethoxyphenyl 2-trifluoromethoxybenzene-                            sulfonyl chloride                                                            2,5-dichlorophenyl 2,5-dichlorobenzenesulfonyl                                 chloride                                                                     2,5-dimethoxy 2,5-dimethoxybenzenesulfonyl                                     chloride                                                                     2-fluorophenyl 2-fluorobenzenesulfonyl chloride                               3-fluorophenyl 3-fluorobenzenesulfonyl chloride                             ______________________________________                                    

Example 90

General Procedure for the Preparation of Compounds of the PresentInvention

Following the four-step protocol outlined in Examples 7 to 10(hydrolysis, coupling, hydrogenation and hydrolysis), certain of theintermediates of Example 89 are used to synthesize the followingcompounds of the present invention (as their trifluoroacetic acidsalts): ##STR117##

(3-phenylsulfonylamino-2-oxo-1,2-dihydro-pyridyl)acetyl-L-argininal(Compound B), ##STR118##

[3-(1-naphthyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR119##

[3-(2-naphthyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR120##

[3-(2-carbomethoxyphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR121##

[3-(2-trifluoromethylbenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR122##

(3-cyclohexylaminosulfonylamino-2-oxo-1,2-dihydro-pyridyl)acetyl-L-argininal,##STR123##

[3-(2-trifluoromethylphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR124##

[3-(3-trifluoromethylphenyl)sulfonylamnino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR125##

[3-(4-trifluoromethylphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR126##

[3-(2-methylphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR127##

[3-(3-methylphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR128##

[3-(2-methyl-5-fluorophenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR129##

[3-(2-methoxylphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,##STR130##

[3-(3-methoxylphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal,and ##STR131##

[3-(2-aminophenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal.

Example 91

Preparation of t-butyl [(t-butyl3-carboxyacetate)-6-methyl-2-oxo-1,2-dihydro-1-pyridyl]acetate##STR132##

To a stirred solution of 2-hydroxy-6-methylpyridine-3-carboxylic acid(12.00 g, 78 mmole) in dimethylformamide (180 ml) was added potassiumcarbonate (22.8 g, 165 mmole) and t-butyl bromoacetate (24.2 mL, 165mmole). After stirring for 36 hours, the reaction mixture was dilutedwith water (700 mL) and extracted with ethyl acetate (2×200 mL). Thecombined organic extracts were washed with brine and dried overmagnesium sulfate. The solvent was removed in vacuo. The residue wassuction chromatographed through flash silica gel using 10-50% ethylacetate/hexanes to yield 22.46 g (75%) of the title compound as an oil.Rf=0.10 (silica gel, 33% ethyl acetate/hexanes).

Example 92

Preparation of t-butyl(3-carboxy-6-methyl-2-oxo-1,2-dihydro-1-pyridyl)acetate ##STR133##

To a stirred solution of the compound of Example 91 (22.46 g, 59 mmole)in tetrahydrofuran (270 mL) was added 1.0M lithium hydroxide (90 mL, 90mmole). After 2 hours, the solution was concentrated. The solution wasdiluted with water (150 mL) and extracted with diethyl ether. Theaqueous layer was acidified to pH 3 with 1M sodium bisulfate, andextracted with ethyl acetate twice. The combined extracts were washedwith brine, dried over magnesium sulfate and the solvent was removed invacuo. The title compound was isolated (18.02 g) in quantitative yield.

Example 93

Preparation of t-butyl(3-benzyloxycarbonylamino-6-methyl-2-oxo-1,2-dihydro-1-pyridyl)acetate##STR134## To the compound of Example 92 (13.0 g, 48.7 mmole) suspendedin dioxane (150 ml), was added triethylamine (7.7 mL, 55 mmole) dropwiserapidly with stirring followed by diphenylphosphoryl azide (16 mL, 73mmole). The suspension was heated for 2 hours using a preheated 110° C.oil bath. Benzyl alcohol (7.6 g, 73 mmole) was then added and themixture was stirred at 110° C. for 20 hours. The reaction mixture wascooled and concentrated. The residue was suspended in ethyl acetate (400mL) and was washed with 3% HCl, then brine, dried over magnesium sulfateand concentrated. The crude product was chromatographed on flash silicagel using 20-67% ethyl acetate/hexanes to afford the title compound(14.2 g, 78% yield) as a white solid. Rf=0.53 (silica gel, 33% ethylacetate/hexanes).

Example 94

Preparation of t-butyl(3-amino-6-methyl-2-oxo-1,2-dihydro-1-pyridyl)acetate ##STR135##

Using similar procedures to that described in Example 106 herein, thetitle compound (0.89 g) was prepared from the compound of Example 93(1.7 g) in 82% yield. Rf=0.69 (silica gel, 10%methanol/dichloromethane).

Example 95

Preparation of t-butyl(3-benzylsufonylamino-6-methyl-2-oxo-1,2-dihydro-1-pyridyl)acetate##STR136##

Collidine (0.59 mL, 4.5 mmole) was added in one portion to a stirredsolution of the compound of Example 94 (0.89 g, 3.7 mmole) andbenzylsulfonyl chloride (0.86 g, 4.5 mmole) in acetonitrile (20 ml)cooled in an ice bath. The solution was stirred for 5 minutes at 0° C.,followed by 45 minutes at room temperature. The reaction mixture wasquenched with water, then diluted with ethyl acetate (100 mL), washedwith 3% HCl (until aqueous layer was pH 1), and brine, dried overmagnesium sulfate, and the solvent was removed. The residue wasdissolved in methanol, concentrated to a volume of approximately 3 mL,and the product was precipitated with the addition of diethyl ether. Theprecipitate was filtered to give 0.67 g of the title compound. Thefiltrate was concentrated and chromatographed on flash silica gel using20 to 67% ethyl acetate hexanes as eluent. An additional 0.20 g of thetitle compound was recovered. A total of 0.87 g of the title compound(59% yield) was recovered. Rf=0.29 (silica gel, 33% ethylacetate/hexanes).

Example 96

Preparation of(3-benzylsulfonylamino-6-methyl-2-oxo-1,2-dihydro-1-pyridyl)acetic acid##STR137##

To a cooled (0° C.) solution of the compound of example 95 (0.87 g, 2.2mmole) in dichloromethane (10 mL) was added trifluoroacetic acid (10mL). After stirring for 30 minutes, the ice bath was removed and thesolution stirred for 2.5 hours at room temperature. The reaction mixturewas concentrated. The resulting solid was triturated with diethyl ether(15 mL) and dried in vacuo. The title compound (0.73 g) was isolated in98% yield. Rf=0.13 (silica gel, 10% methanol/dichloromethane).

Example 97

Preparation of 2-hydroxy-6-ethylpyridine-3-carbonitrile ##STR138##

To a suspension of 1-hydroxy-6-methylpyridine-3-carbonitrile (12.24 g,0.091 mole) in tetrahydrofuran (100 mL) cooled to -78° C. under anitrogen atmosphere was added dropwise lithium diisopropylamide (100 mLof a 2.0M solution in heptane/tetrahydrofuran/ethylbenzene, 0.20 mole).After the addition was complete, the solution was stirred in an ice bathfor 2 hours. Iodomethane (6.25 mL, 0.10 mol) was added, and the reactionmixture was stirred for an additional 2.5 hours at 0° C. and 30 minutesat room temperature. Water (300 mL) and 1.0N NaOH 50 mL) were added. Theaqueous solution was washed with ethyl acetate (150 mL), acidified with1.0N sodium bisulfate to pH 4, and extracted with 10% methanol/ethylacetate twice (500 mL total). Sodium chloride was added to the aqueouslayer, and the solution was extracted with 10% isopropanol/ethyl acetatetwice (500 mL total). The combined organic layers were washed withbrine, dried over magnesium sulfate and the solvent removed underreduced pressure. The residue was recrystallized frommethanol/isopropanol to give the title compound (7.56 g) as orangeneedles in 56% yield. Rf=0.26 (silica gel, 10% isopropanol/chloroform);m.p. 235 to 240° C. (decomp).

Example 98

Preparation of 2-hydroxy-6-ethylpyridine-3-carboxylic acid ##STR139##The compound of Example 97 (7.56 g, 51 mmole) was refluxed in 50%sulfuric acid (50 mL) for 3 hours. The reaction mixture was cooled, andpoured into water (250 mL). The solution was allowed to stand at 4° C.for 16 hours. The solid was filtered, washed with water and air dried toafford the title compound (6.03 g) in 71% yield as a tan solid; m.p.190.5 to 193° C.

Example 99

Preparation of3-benzyloxycarbonylamino-6-ethyl-2-oxo-1,2-dihydro-1-pyridine ##STR140##

Using similar procedures to that described above in Example 93, thetitle compound (0.74 g) was prepared from the compound of Example 98(1.00 g) in 45% yield. Rf=0.18 (silica gel, 20% ethylacetate/dichloromethane); m.p. 153.5 to 154° C.

Example 100

Preparation of t-butyl(3-benzyloxycarbonylamino-6-ethyl-2-oxo-1,2-dihydro-1-pyridyl)acetate##STR141##

To a solution of the compound of Example 99 (150 mg, 0.55 mmole) intetrahydrofuran (2 mL) was added lithium hexamethyldisilazide (0.61 mLof a 1.0 M solution in tetrahydrofuran, 0.61 mmole). After 1.5 hours,t-butyl bromoacetate (0.089 mL, 0.61 mmole) was added. The reactionmixture was stirred for 16 hours, then diluted with water (5 mL) andsaturated ammonium chloride (5 mL), and extracted with ethyl acetate(3×5 mL). The combined organic layers were washed with brine, dried overmagnesium sulfate and the solvent was reduced. Hexanes (10 mL) wereadded, and the solvent was removed in vacuo to afford 0.20 g of thetitle compound as a white solid in 94% yield. Rf=0.76 (silica gel, 20%ethyl acetate/dichloromethane).

Example 101

Preparation of(3-benzylsulfonylamino-6-ethyl-2-oxo-1,2-dihydro-1-pyridyl)acetic acid

Following the three step protocol outlined in Examples 94-96, theintermediate of Example 100 is used to synthesize the following compoundof the present invention. ##STR142##

Example 102

Ethyl 2-methyl-pyrimidin-6(1H)-one-5-carboxylate ##STR143##

Acetamidine acetate (37.21 g, 0.31 mole) and diethylethoxymethylenemalonate (63 mL, 0.31 mole) were refluxed for 4 h inethanol (60 mL). The reaction mixture was allowed to cool for 15minutes, then acetamidine acetate (37.21 g, 0.31 mole) was added. Thereaction mixture was refluxed for 22 hours, allowed to cool to roomtemperature, and diluted with water (200 mL) and dichloromethane (200mL). The aqueous layer was extracted with 10%isopropanol/dichloromethane (2×200 mL). The combined organic extractswere washed with water (50 mL), brine (50 mL), dried over magnesiumsulfate, filtered and the solvent was removed. The residue wasrecrystallized from chloroform/hexanes in two crops to afford the titlecompound (24.92 g) in 46% yield as yellowish crystals. R_(f) =0.27(silica gel, 10% isopropanol in dichloromethane); m.p. 187 to 188° C.

Example 103

Preparation of Ethyl 3-(t-butylacetyl)-2-methyl-pyrimidin-6(1H)-one-5-carboxylate ##STR144##

Tetra-n-butylammonium fluoride(27.4 mL of a 1.0 M solution intetrahydrofuran, 27.4 mmole) was diluted with hexanes (30 mL), thesolvent was removed under reduced pressure, and the white crystals weretaken up in dimethoxyethane (50 mL). t-Butyl bromoacetate (3.0 mL, 20.1mmole) was added while stirring, followed by the compound of Example 102(2.50 g, 13.7 mmole). The mixture was stirred under a nitrogenatrmospherre for 1.5 hours at room temperature. The reaction mixture wasdiluted with water (100 mL), and extracted with ethyl acetate (3×50 mL).The combined organic extracts were washed with water, then brine (20 mLeach), dried over magnesium sulfate, and the solvent was removed. Theresidue was purified through flash silica gel using 50% ethyl acetate,then ethyl acetate as eluent. The title compound was isolated to yield1.81 g (45%). Rf=0.24 (silica gel, 20% ethyl acetate indichloromethane).

Example 104

Preparation of 3-(t-butylacetyl)-2-methyl-pyrimidin-6(1H)-one-5-carboxylic acid ##STR145##

To the compound of Example 103 (10.16 g, 0.034 mole), suspended inmethanol (70 ml) and cooled in an ice bath, 1.0N lithium hydroxide (38mL, 0.038 mole) was added dropwise rapidly with stirring. The ice bathwas removed. After 2 h, reaction mixture was neutralized to pH 7 with1.0N hydrochloric acid. The solvent was reduced under vacuum, theresidue diluted with water (50 mL) and washed with ethyl acetate (2×25mL) The aqueous layer was acidified with 2.3N HCl to pH 1, extractedwith ethyl acetate (50 mL), followed by dichloromethane twice (30 mLtotal). The combined organic extracts are washed with brine (3×10 mL).The solvent was dried over magnsium sulfate, and removed in vacuo. Theresidue was recrystallized from ethyl acetate/diethyl ether (first crop)and ethyl acetate/diethyl ether/hexanes (second crop) to afford 4.37 g(48%) of the title compound. Rf=0.31 (silica gel, 1% acetic acid/10%isopropanol in chloroform).

Example 105

Preparation of t-butyl2-methyl-5-benzyloxycarbonylamino-6-oxo-1,6-dihydro-1-pyrimidinylacetate##STR146##

To the compound of Example 104 (4.20 g, 0.0157 mole) suspended indioxane (50 ml), was added triethylamine (4.4 mL, 0.0313 mole) dropwiserapidly with stirring followed by diphenylphosphoryl azide (3.7 mL,0.0172 mole). The suspension was heated for 2 hours using a preheated100° C. oil bath. Benzyl alcohol (3.2 g, 0.0313 mole) was then added andthe mixture was stirred at 100° C. overnight. The reaction mixture wascooled and concentrated. The residue was suspended in ethyl acetate (100mL) and was washed with saturated ammonium chloride, 1.0N NaOH, water(twice) and brine. The extract was dried over magnesium sulfate andconcentrated. The crude product was chromatographed on flash silica gelusing 10 to 25% ethyl acetate/dichloromethane to give the title compound(3.07 g, 53% yield) as a light yellow solid. R_(f) =0.24 (silica gel,20% ethyl acetate in dichloromethane).

Example 106

Preparation of t-butyl2-methyl-5-amino-6-oxo-1,6-dihydro-1-pyrimidinylacetate ##STR147##

The compound of Example 105 (1.50 g, 4.0 mmol) was hydrogenated over 10%palladium on carbon (0.16 g) in ethanol (30 mL) at balloon pressureovernight. Celite was added, and the solution was filtered. The solventwas reduced. Hexanes were added and the solvent was removed in vacuo toafford 0.97 g (quantitative yield) of the title compound as a whitesolid.

R_(f) =0.24 (silica gel, 10% isopropanol in chloroform).

Example 107

Preparation of t-butyl2-methyl-5-benzylcarbonyloxyamino-6-oxo-1,6-dihydro-1-pyrimidinylacetate##STR148##

Benzylsulfonyl chloride (1.06 g, 5.6 mmole) was added in one portion toa stirred solution of the compound of example 106 (0.89 g, 3.7 mmole)and 4-methylmorpholine (1.47 mL, 11.2 mmole) in tetrahydrofuran (10 ml).The solution was stirred for 2 hours. The reaction mixture wasconcentrated, then diluted with ethyl acetate (100 mL), washed with 1.0NHCl (until aqueous layer is pH 1), water, saturated sodium bicarbonateand brine. The organic layer was dried over magnesium sulfate, and thesolvent was removed. The residue was recrystallized from ethyl acetate(first crop) and ethyl acetate/ether/hexanes (second crop). The secondcrop was treated with 1.0 M potassium carbonate (3 mL) and methanol(10mL) for 2 hours. The solution became homogeneous. The reaction mixturewas acidified to pH 7 with 1.0N HCl. The solvent was reduced, and theaqueous solution was extracted with ethyl acetate three times. Thecombined organic extracts were washed with brine, dried over magnesiumsulfate and the solvent was removed in vacuo. In a similar fashion thefirst crop was treated with 1.0M potassium carbonate and methanol. Atotal of 1.20 g of the title compound (82% yield) was recovered as awhite solid. R_(f) =0.26 (silica gel, 20% ethyl acetate indichloromethane).

Example 108

Preparation of2-methyl-5-benzylsulfonylamino-6-oxo-1,6-dihydro-1-pyrimidinylaceticacid ##STR149##

To the compound of Example 107 (1.15 g, 2.9 mmole) was treated with 50%trifluoroacetic acid/dichloromethane (10 mL). After 1 hour, the reactionmixture was concentrated, then diluted with diethyl ether (100 mL). Thesolution was allowed to stand overnight, then the solvent was removedunder reduced pressure. The residue was partitioned between saturatedsodium bicarbonate (25 mL) and ethyl acetate (10 mL). The aqueous layerwas washed with ethyl acetate, then acidified with 2.3M HCl to pH 1. Thepreciptiate was filtered, washed with water and dried under vacuum togive the title compound (0.75 g, 76% yield) as a white solid. m.p. 244to 246° C. (decomp.).

Example 109

Preparation of 5-nitro-1-methyl-uracil ##STR150##

5-nitrouracil (10.00 g, 64 mmole) and potassium carbonate were stirredin dimethylformamide (50 mL) for 15 min. A solid formed. Iodomethane(5.3 mL, 85 mmol) was added and the flask was shaken until the soliddissolved. After the reaction mixture was stirred for 30 min., 2% NaOH(w/v) (200 mL) was added, followed by water (100 mL). The solution waswashed with ethyl acetate (100 mL), and the aqueous layer was acidifiedto pH 3 with 1.0N HCl. A precipitate formed as the pH was lowered. Afterallowing the heterogeneous solution to stand for 16 hours, the productwas filtered, washed with water, and air dried. The title compound wasisolated in 77% yield as a yellow powder; m.p. 249 to 250° C.

Example 110

Preparation of t-butyl (5-nitro-1-methyl-uracilyl)acetate ##STR151##

Sodium hydride (0.51 g of a 60% dispersion in mineral oil, 13 mmol) waswashed with pentane three times (4 mL each). The compound of Example 109(2.00 g, 12 mmole) was added portionwise. After the addition wascomplete, the reaction mixture was stirred for 30 minutes under anitrogen atmosphere. t-Butyl bromoacetate (1.73 g, 12 mmole) was addedin one portion, and the solution was stirred for 3 hours. The reactionmixture was diluted with water (200 mL), and extracted with ethylacetate (3×50 mL). The combined organic extracts were washed with water(3×50 mL), then brine, dried over magnesium sulfate. The solvent wasconcentrated, hexanes were added and the solvent was removed in vacuo toafford the title compound (1.97 g) in 59% yield. Rf=0.38 (silica gel,20% ethyl acetate in dichloromethane.

Example 111

Preparation of Ethyl 5-(benzylsulfonylamino-1-methyl-uracilyl)acetate##STR152##

Using similar procedures to that described above in Example 6, butemploying 3 equivalents of 4-methyl morpholine as base during thereaction with benzylsulfonyl chloride, the title compound was preparedfrom the compound of Example 110 in 48% yield; m.p. 165 to 166° C.

Example 112

Preparation of 5-(benzylsulfonylamino-1-methyl-uracilyl)acetic acid##STR153##

Using a similar procedure to that described above in Example 108, thetitle compound was prepared from the compound of Example 111 in 88%yield; m.p. 200 to 201° C.

Example 113

General Procedure for Preparation of Compounds of the Present Invention

Following the three-step protocol outlined in Examples 8 to 10(coupling, hydrogenation, hydrolysis) the intermediates of Examples 96,108 and 112 were used to synthesize the following compounds of thepresent invention: ##STR154##

(3-benzylsulfonylamino-6-methyl-2-oxo-1,2-dihydro-1-pyridyl)acetyl-L-argininal,trifluoroacetate salt (Compound C); ##STR155##

(5-benzylsulfonylamino-2-methyl-6-oxo-1,6-dihydro-1-pyrimidinyl)acetyl-L-argininal, trifluoroacetate salt(Compound D); and ##STR156##

(5-benzylsulfonylamino-1-methyl-uracilyl)acetyl-L-argininal,trifluoroacetate salt (Compound E).

Example 114

Preparation of4-(2-trimethylsiloxyphenethyl)-3-nitro-2-oxo-1,2-dihydro-1-pyridine##STR157##

A suspension of 4-methyl-3-nitro-2-pyridone (3.08 g, 20 mmole) intetrahydrofuran (50 mL) was cooled to 0° C. Lithium hexamethyldisilazide(21 mL of a 1M solution in tetrahydrofuran, 21 mmole) was added to thereaction over 15 minutes. After stirring for 45 minutes, trimethylsilylchloride (2.7 mL, 21 mmole) was added. After an hour, another portion oflithium hexamethyldisilazide (21 mL of a 1M solution in tetrahydrofuran,21 mmole) was added to the solution. After 30 minutes, freshly distilledbenzaldehyde (2.1 mL, 21 mmole) was added. The reaction was allowed towarm to room temperature and after 18 hours, it was quenched withaqueous ammonium chloride (20 ml), extracted with ethyl acetate (150ml), washed with brine (50 ml), and dried over magnesium sulfate. Theproduct was purified by chromatography with silica gel, eluting with2-10% methanol/dichloromethane. Recrystallization from toluene yielded0.79 g (8.5%) of the title compound. R_(f) =0.25 (silica gel, 50% ethylacetate/hexanes).

Example 115

Preparation of Ethyl[4-(2-trimethylsiloxyphenethyl)-3-nitro-2-oxo-1,2-dihydro-1-pyridyl]acetat##STR158##

To a solution of the compound of Example 114 (0.79 g, 2.4 mmole) intetrahydrofuran at 0° C. was added lithium hexamethyldisilazide (2.5 mLof a 1M in tetrahydrofuran. 2.5 mmole) over 5 minutes. After 30 minutes,ethyl bromoacetate (0.28 mL, 2.5 mmole) was added. The reaction wasallowed to warm to room temperature and after 8 hours, it was quenchedwith aqueous ammonium chloride (5 mL), extracted with ethyl acetate (75mL), washed with brine (30 mL), and dried over magnesium sulfate. Theproduct was purified by chromatography with silica gel, eluting with(33%) ethyl acetate/hexanes to yield 0.76 g of the title compound (81%yield). R_(f) =0.45 (silica gel, 50% ethyl acetate/hexanes).

Example 116

Preparation of[3-acetamido-4-(2-hydroxyphenethyl)-2-oxo-1,2-dihydro-1-pyridyl]aceticacid ##STR159##

The compound of Example 115 (760 mg, 1.82 mmole) was dissolved in ethylacetate (10 mL). Acetic anhydride (0.69 mL, 7.3 mmole) and 10% palladiumon carbon (75 mg) were added and the reaction was stirred under ahydrogen balloon for 18 hours. The reaction was then filtered throughcelite and concentrated. The residue was dissolved in tetrahydrofuran(7.0 mL) and 1.0M lithium hydroxide was added (3.6 mL, 3.6 mmole). Thereaction was stirred for 22 hours at which time additional 1.0M lithiumhydroxide (2.0 mL, 2.0 mmole) and methanol (2.0 mL) were added. Thereaction was stirred for 48 hours at room temperature, then was dilutedwith water (20 mL) and washed with ethyl acetate (20 mL). The aqueouslayer was acidified with concentrated HCl to pH˜3, and the product wasextracted into ethyl acetate (50 mL), washed with brine (40 mL), anddried over magnesium sulfate. Back extraction of the aqueous layer with30% isopropanol/dichloromethane yielded a total of 440 mg (96%) of thetitle compound. ¹ H NMR (CD₃ OD): delta 2.15 (3H, s), 2.81-2.94 (2H, m),4.70 (2H, s), 4.9 (1H, dd, J=5.3, 8.2 Hz), 6.3 (1H, d, J=7.1 Hz),7.22-7.36 (5H, m), 1.73 (1H, d, J=7.1 Hz).

Example 117

General Procedure for Preparation of Compounds of the Present Invention

Following the three-step protocol outlined in Examples 8 to 10(coupling, hydrogenation, hydrolysis) the intermediates of Examples 101and 116 are used to synthesize the following compounds of the presentinvention: ##STR160##

(3-benzylsulfonylamino-6-ethyl-2-oxo-1,2-dihydro-1-pyridyl)acetyl-L-argininal,trifluoroacetate salt, and ##STR161##

[3-acetamido-4-(2-hydroxyphenethyl)-2-oxo-1,2-dihydro-1-pyridyl]acetyl-L-argininal,trifluoroacetate salt.

Example 118

(5-chloro-2-methoxy-phenylsulfonyl-3-amino-2-oxo-1,2-dihydropyridyl)acetyl-L-N^(g)-nitro-argininal-ethyl cyclol ##STR162##

Following the two-step protocol outlined in Examples 7 to 8 (hydrolysis,coupling), the title compound is prepared from an intermediate ofExample 89, ethyl(2-methoxy-5-chloro-benzenesulfonyl-3-amino-2-oxo-1,2-dihydropyridyl)acetate.

Example 119

Preparation of(5-chloro-2-methoxy-phenylsulfonyl-3-amino-2-oxo-1,2-dihydropyridyl)acetyl-L-Argininal,trifluoroacetate salt ##STR163##

To a stirred solution of the compound of Example 118 (280 mg, 0.47 mmol)in ethyl alcohol (5 mL), a mixture of freshly made 20% titanium(III)chloride solution in water (3.7 mL, 4.7 mmol) and 4.0M ammonium acetatebuffer, pH 5.0 (7.4 mL) was added. The reaction mixture was stirred atroom temperature. After the reaction was complete (30-45 min.), theexcess of titanium(III) chloride was oxidized by bubbling the airthrough the reaction mixture (30 min.). The solvent was removed invacuo. The residual was taken into water (50 mL) and then centrifuged at3,000 rpm for 10 minutes. The supernatant was decanted, and the solidwas washed with water (30 mL) and centrifuged. The combined supernatantswere concentrated to 25 mL. The solution was cooled down to 0° C. withice bath. 12N hydrochloric acid (25 mL) was added, and the ice bath wasremoved. The reaction mixture was stirred at room temperature. After thereaction was complete (30-45 minutes), the reaction mixture was quenchedwith water (150 mL) and sodium acetate (40 g), and then filtered. Theaqueous solution was purified by reverse phase HPLC with C-18 columnusing a gradient system of 17 to 35% acetonitrile in water with 0.1% oftrifluoroacetic acid over 30 min. to afford 160 mg of the title compound(160 mg, 0.31 mmol). MS: 513 (M+H⁺).

Example 120

General Procedure for Reaction of Ethyl(3-amino-2-oxo-1,2-dihydropyridyl)acetate with sufonyl chlorides

To a stirred solution of ethyl (3-amino-2-oxo-1,2-dihydropyridyl)acetate (5.89 g, 30 mmole) in dry tetrahydrofuran (300 mL) is added2,4,6-collidine (7.93 mL, 60 mmol) and the solution is cooled to 0° C.under nitrogen. The appropriate sulfonyl or sulfamoyl chloride listedbelow (33 mmol) dissolved in tetrahydrofuran (25 to 75 mL) is addeddropwise. After the addition is complete, the reaction mixture isstirred for 30 minutes to 1 hour at 0° C. and then at ambienttemperature for 0 to 72 hours. The reaction mixture is diluted withethyl acetate, washed successively with 1.0N HCl, water, saturatedsodium bicarbonate and brine, dried over magnesium sulfate, and thesolvent removed in vacuo. The residue is chromatographed on silica gelusing a gradient system of dichloromethane and 1 to 4% methanol indichloromethane to afford the product, judged pure by TLC (silica gel).Using this method and the starting materials listed below, intermediateshaving the formula given below are made:

    ______________________________________                                          #STR164##                                                                   R =             Starting material                                             ______________________________________                                        2-fluorophenyl  2-fluorobenzenesulfonyl chloride                                3-fluorophenyl 3-fluorobenzenesulfonyl chloride                               2-trifluoromethoxyphenyl 2-trifluoromethoxybenzene-                            sulfonyl chloride                                                            2,5-dimethylphenyl 2,5-dimethylbenzenesulfonyl                                 chloride                                                                     2,5-dimethoxyphenyl 2,5-dimethoxybenzenesulfonyl                               chloride                                                                     2,6-difluorophenyl 2,6-difluorobenzenesulfonyl                                 chloride                                                                     phenethyl 2-phenylethanesulfonyl chloride                                     cyclohexylmethyl cyclohexylmethanesulfonyl                                     chloride                                                                     2,5-dichlorophenyl 2,5-dichlorobenzenesulfonyl                                 chloride                                                                     2-fluorobenzyl (2-fluorophenyl)methanesulfonyl                                 chloride                                                                     3-fluorobenzyl (3-fluorophenyl)methanesulfonyl                                 chloride                                                                     3-trifluoromethylbenzyl (3-trifluoromethyl-                                    phenyl)methanesulfonyl chloride                                              2-carbomethoxybenzyl (2-carbomethoxyphenyl)methane-                            sulfonyl chloride                                                            3-carbomethoxybenzyl (3-carbomethoxyphenyl)methane-                            sulfonyl chloride                                                            2,6-difluorobenzyl (2,6-difluorophenyl)methane-                                sulfonyl chloride                                                            2,5-difluorobenzyl (2,5-difluorophenyl)methane-                                sulfonyl chloride                                                            2,4-difluorobenzyl (2,4-difluorophenyl)methane-                                sulfonyl chloride                                                            2-carbomethoxy-5- 2-carbomethoxy-5-fluorobenzyl                               fluorobenzyl sulfonyl chloride(see Example                                    148)                                                                          2-carbomethoxy-6- 2-carbomethoxy-6-fluorobenzyl                               fluorobenzyl sulfonyl chloride (see Example                                   149)                                                                          4-methoxyphenyl 4-methoxyphenylsulfonyl chloride                              3,4-dimethoxyphenyl 3,4-dimethoxyphenylsulfonyl                               chloride                                                                    ______________________________________                                    

Example 121

General Procedure for the Preparation of Compounds of the PresentInvention

Following the four-step protocol outlined in Examples 7 to 10(hydrolysis, coupling, hydrogenation and hydrolysis), the intermediatesof Example 120 are used to synthesize the following compounds of thepresent invention (as their trifluoroacetic acid salts): ##STR165##

3-(2-fluorophenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal (Compound121A); ##STR166##

3-(3-fluorophenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121B); ##STR167##

(3-(2-trifluoromethoxyphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl)acetyl-L-argininal(Compound 121C); ##STR168##

[3-(2,5-dimethylphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121D); ##STR169##

[3-(2,5-dimethoxyphenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121E); ##STR170##

[3-(2,6-difluorophenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121F); ##STR171##

[3-(phenethyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121G); ##STR172##

(3-cyclohexylmethylsulfonylamino-2-oxo-1,2-dihydro-pyridyl)acetyl-L-argininal(Compound 121H); ##STR173##

[3-(2,5-dichlorophenyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121I); ##STR174##

3-(2-fluorobenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121J); ##STR175##

3-(3-fluorobenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121K); ##STR176##

3-(3-trifluoromethylbenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121L); ##STR177##

3-(2-carbomethoxybenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121M); ##STR178##

3-(3-carbomethoxybenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121N); ##STR179##

[3-(2,6-difluorobenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 1210); ##STR180##

[3-(2,5-difluorobenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121P); ##STR181##

[3-(2,4-difluorobenzyl)sulfonylamino-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 121Q); ##STR182##

[3-(2-carbomethoxy-5-fluorobenzyl)sulfonylamino-2-oxo-1,2-dihydropyridyl]acetyl-arginine(Compound 121R); ##STR183##

[3-(2-carbomethoxy-6-fluorobenzyl)sulfonylamino-2-oxo-1,2-dihydropyridyl]acetyl-arginine(Compound 121S); ##STR184##

[3-(4-dimethoxyphenyl)sulfonylamino-2-oxo-1,2-dihydropyridyl]acetyl-arginine(Compound 121T); and ##STR185##

[3-(3, 4-dimethoxyphenyl)sulfonylamino-2-oxo-1,2-dihydropyridyl]acetyl-arginine (Compound 121U).

Correct molecular weight for the compounds is confirmed by massspectroscopy.

Example 122

Preparation of t-Butyl(3-amino-6-ethyl-2-oxo-1,2-dihydro-1-pyridyl)acetate ##STR186##

Using similar procedures to that described in Example 106 herein, thetitle compound (5.79 g) was prepared from the compound of Example 100(8.12 g) in quantitative yield. R_(f) =0.05 (silica gel, 33% ethylacetate/hexanes).

Example 123

Preparation of t-Butyl(3-amino-6-ethyl-2-oxo-1,2-dihydro-1-pyridyl)acetate ##STR187##

Following the four step protocol as outlined in Examples 97 to 100,substituting 2-bromopropane (10.3 mL, 0.10 mol) for iodomethane, thetitle compound is prepared. Rf=0.69, (silica gel, 50% ethylacetate/hexanes).

Example 124

General Procedure for Reaction of t-Butyl(3-amino-6-alkyl-2-oxo-1,2-dihydro-1-pyridyl)acetate with sulfonylchlorides

To a stirred solution of the compound of Example 94 (7.15 g, 30 mmole),the compound of Example 122 (7.60 g, 30 mmole), OR the compound ofExample 123 (8.50 g, 30 mmole), in dry tetrahydrofuran (300 mL) is added2,4,6-collidine (7.93 mL, 60 mmole) and the solution is cooled to 0° C.under nitrogen. The appropriate sulfonyl chloride, listed below, (33mmole) dissolved in tetrahydrofuran (25 to 75 mL) is added dropwise.After the addition is complete, the reaction mixture is stirred for 30minutes to 1 hour at 0° C. and then at ambient temperature for 0 to 72hours. The reaction mixture is diluted with ethyl acetate, washedsuccessively with 1.0N HCl, water, saturated sodium bicarbonate andbrine, dried over magnesium sulfate, and the solvent is removed invacuo. The residue is chromatographed on silica gel using a gradientsystem of dichloromethane and 1 to 4% methanol in dichloromethane toafford the product, judged pure by TLC (silica gel). Using this methodand the starting materials listed below, intermediates having theformula given below are made:

    ______________________________________                                          #STR188##                                                                       Product Compound   Starting materials                                     ______________________________________                                        R.sub.1 = 2-trifluoromethylbenzyl,                                                               Compound of Example 94,                                      R.sub.2 = CH.sub.3 (2-trifluoromethyl-                                         phenyl)methanesulfonyl                                                        chloride                                                                     R.sub.1 = 2-methyl-5-fluorophenyl, Compound of Example 94,                    R.sub.2 = CH.sub.3 2-methyl-5-fluorobenzene-                                   sulfonyl chloride                                                            R.sub.1 = 2,5-dimethoxyphenyl,, Compound of Example 94,                       R.sub.2 = CH.sub.3 2,5-dimethoxybenzene-                                       sulfonyl chloride                                                            R.sub.1 = 2-carbomethoxybenzyl, Compound of Example 94,                       R.sub.2 = CH.sub.3 2,6-difluorobenzenesulfonyl                                 chloride                                                                     R.sub.1 = benzyl, Compound of Example 122,                                    R.sub.2 = CH.sub.2 CH.sub.3 benzylsulfonyl chloride                           R.sub.1 = 2-methyl-5-fluorophenyl, Compound of Example 122,                   R.sub.2 = CH.sub.2 CH.sub.3 2-methyl-5-fluorobenzene-                          sulfonyl chloride                                                            R.sub.1 = benzyl, R.sub.2 = CH.sub.2 CH(CH.sub.3).sub.2 Compound of                            Example 123,                                                  benzylsulfonyl chloride                                                    ______________________________________                                    

Example 125

General Procedure for Reaction of(3-amino-6-alkyl-2-oxo-1,2-dihydropyridyl) acetic acid withtrifluoroacetic acid

The intermediates of Example 124 (3.0 mmole) are treated with 50%trifluoroacetic acid/dichloromethane (10 mL). After 1 hour, the reactionmixture is concentrated, then diluted with toluene. The solution isagain concentrated, toluene is added, and the solvent is removed invacuo. Using this method and the starting materials listed below,intermediates having the formula given below are made: ##STR189##

R₁ ═2-trifluoromethylbenzyl, R₂ ═CH₃ ;

R₁ ═2-methyl-5-fluorophenyl, R₂ ═CH₃ ;

R₁ ═2,5-dimethoxyphenyl, R₂ ═CH₃ ;

R₁ =2-carbomethoxybenzyl, R₂ ═CH₃ ;

R₁ =benzyl, R₂ ═CH₂ CH₃ ;

R₁ ═2-methyl-5-fluorophenyl, R₂ ═CH₂ CH₃ ;

R₁ =benzyl, R₂ ═CH₂ CH(CH₃)₂ ;

R₁ ═2-fluorobenzyl, R₂ ═CH₃ ; and

R₁ ═2-carbomethoxy-5-fluorobenzyl, R₂ ═CH₃.

Example 126

General Procedure for the Preparation of Compounds of the PresentInvention:

Following the three-step protocol outlined in Examples 8 to 10(coupling, hydrogenation and hydrolysis), the intermediates of Example125 were used to synthesize the following compounds of the presentinvention (as their trifluoroacetic acid salts): ##STR190##

[3-(2-trifluoromethylbenzyl)sulfonylamino-6-methyl-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 126A); ##STR191##

[3-(5-fluoro-2-methylphenyl)sulfonylamino-6-methyl-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 126B); ##STR192##

3-[(2,5-dimethoxyphenyl)sulfonylamino-6-methyl-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 126C); ##STR193##

[3-(2-carbomethoxybenzyl)sulfonylamino-6-methyl-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 126D); ##STR194##

(3-benzylsulfonylamino-6-ethyl-2-oxo-1,2-dihydro-pyridyl)acetyl-L-argininal(Compound 126E); ##STR195##

[3-(5-fluoro-2-methylphenyl)sulfonylamino-6-ethyl-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 126F); ##STR196##

(3-benzylsulfonylamino-6-isobutyl-2-oxo-1,2-dihydro-pyridyl)acetyl-L-argininal(Compound 126G); ##STR197##

[3-(2-fluorobenzyl)sulfonylamino-6-methyl-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 126H); and ##STR198##

[3-(2-carbomethoxy-5-fluorobenzyl)sulfonylamino-6-methyl-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 126I).

Example 127

Preparation of t-butyl [3-nitro-2-oxo-1,2-dihydropyridyl]acetate##STR199##

Using similar procedures to those described in Example 5 herein, thetitle compound (6.77 g) was prepared from 3-nitro-2-hydroxypyridine(5.00 g) in 74% yield. R_(f) =0.60 (silica gel, 20% ethylacetate/dichloromethane).

Example 128

Preparation of t-Butyl[3-nitro-2-oxo-4-(3-phenylpropyl)-1,2,3,4-tetrahydropyridyl]acetate##STR200##

To a suspension of magnesium turnings (3.00 g, 49.6 mmole) intetrahydrofuran (30 mL) was added (3-bromopropyl)benzene (9.87 g, 49.6mmole) in tetrahydrofuran (20 mL) under a nitrogen atmosphere. Afterapproximately 2 to 3 ml of the (3-bromopropyl)benzene solution wereadded, a few milligrams of iodine were added, and the reaction mixturewas heated gently for 3 minutes. The rest of the (3-bromopropyl)benzenesolution was added over 10 minutes. The solution was stirred in a 50 to60° C. oil bath overnight to give a solution of 3-phenylpropyl magesiumbromide.

A solution of the compound of Example 127 (6.00 g, 23.6 mmol) and zincchloride (6.44 g, 47.2 mmole) in tetrahydrofuran (50 mL) under an argonatmosphere was stirred for 10 minutes, then was cooled in an ice bath.The solution of 3-phenylpropyl magesium bromide from this Example wasadded over a 5 minute period. After stirring 5 minutes at 0° C., the icebath was removed, and the solution was allowed to stir overnight. Thereaction mixture was poured into ethyl acetate (300 mL) and saturatedcitric acid (50 mL). The organic layer was washed with saturated sodiumbicarbonate, water, and then brine (30 mL each). The solution was driedover sodium sulfate, the solvent was removed, and the residue waschromatographed through silica gel using 0 to 60% ethyl acetate/hexanesto afford 6.0 g (68% yield) of the title compound. R_(f) =0.71 (silicagel, 50% ethyl acetate/hexanes).

Example 129

Preparation of t-Butyl[3-nitro-2-oxo-4-(3-phenylpropyl)-1,2-dihydropyridyl]acetate ##STR201##

The compound of Example 128 (6.0 g, 16 mmole) was dissolved intetrahydrofuran (50 mL) under a nitrogen atmosphere. Cesium carbonate(6.3 g, 19.2 mmole, 1.2 equivalents) was added, and the solution wasstirred for 10 minutes. Palladium acetate (3.63 g, 16.2 mmole, 1equivalent) was added, and the solution was stirred overnight. Celiteand silica gel were added, and the reaction mixture was filtered toremove the cesium and palladium salts, washing the solid with ethylacetate (500 mL). The solvent was removed from the filtrate, and theresidue was chromatographed on silca gel using 0 to 50% ethylacetate/hexanes as eluent to afford 2.86 g of the title compound in 48%yield. R_(f) =0.52 (silica gel, 50% ethyl acetate/hexanes).

Example 130

Preparation of t-Butyl[3-amino-2-oxo-4-(3-phenylpropyl)-1,2-dihydropyridyl]acetate ##STR202##

The compound of Example 129 (1.0 g, 2.7 mmole) in methanol (20 mL) washydrogenated under balloon pressure over 10% palladium on carbon (0.15g) for 2 hours. The reaction mixture was filtered, the solvent wasremoved in vacuo to give the title compound (0.92 g) in quantitativeyield. R_(f) =0.42 (silica gel, 50% ethyl acetate/hexanes).

Example 131

Preparation of t-butyl[3-amino-2-oxo-4-(2-phenylethyl)-1,2-dihydropyridyl]acetate ##STR203##

Following the three step protocol, substituting (2-bromoethyl)benzene(6.77 mL, 49.6 mmole) for (3-bromopropyl)benzene, as outlined inExamples 128 to 130, the title compound is prepared. R_(f) =0.63,(silica gel, 10% isopropanol/dichloromethane).

Example 132

General Procedure for Reaction of t-butyl(3-amino-4-alkyl-2-oxo-1,2-dihydropyridyl)acetate with sulfonylchlorides.

To a stirred solution of the compound of Example 130 (10.2 g, 30 mmole)OR the compound of Example 131 (9.78 g, 30 mmole), in drytetrahydrofuran (300 mL) is added the 2,4,6-collidine (7.93 mL, 60mmole) and the solution is cooled to 0° C. under nitrogen. Theappropriate sulfonyl or sulfamoyl chloride listed below (33 to 150mmole) dissolved in tetrahydrofuran (25 to 75 mL) is added dropwise.After the addition is complete, the reaction mixture is stirred for 30minutes to 1 hour at 0° C. and then at ambient temperature for 0 to 72hours. The reaction mixture is diluted with ethyl acetate, washedsuccessively with 1.0N HCl, water, saturated sodium bicarbonate andbrine, dried over magnesium sulfate, and the solvent is removed invacuo. The residue is chromatographed on silica gel using a gradientsystem of dichloromethane and 1 to 4% methanol in dichloromethane toafford the product, judged pure by TLC (silica gel). Using this methodand the starting materials listed below, intermediates having theformula given below are made:

    ______________________________________                                          #STR204##                                                                   Intermediate      Starting materials                                          ______________________________________                                        n = 3, R = methyl Compound of Example 130,                                       methanesulfonyl chloride                                                     n = 2, R = methyl Compound of Example 131,                                     methanesulfonyl chloride                                                     n = 2, R = 2,2,2-trifluoro- Compound of Example 131,                          ethyl 2,2,2-trifluoroethanesulfonyl                                            chloride                                                                     n = 2, R = phenyl Compound of Example 131,                                     benzenesulfonyl chloride                                                     n = 2, R = methylamino Compound of Example 131,                                methylsulfamoyl chloride                                                   ______________________________________                                    

Example 133

General Procedure for Reaction of(3-amino-6-alkyl-2-oxo-1,2-dihydropyridyl)acetic acid withtrifluoroacetic acid

The intermediates of Example 132 (3.0 mmole) are treated with 50%trifluoroacetic acid/dichloromethane (10 mL). After 1 hour, the reactionmixture is concentrated, then diluted with toluene. The solution isconcentrated, toluene is added, and the solvent is removed in vacuo.Using this method and the starting materials listed below, intermediateshaving the formula given below are made: ##STR205##

n=3, R=methyl;

n=2, R=methyl;

n=2, R=2,2,2-trifluoroethyl;

n=2, R=phenyl; and

n=2, R=methylamino.

Example 134

General Procedure for the Preparation of Compounds of the PresentInvention

Following the three-step protocol outlined in Examples 8 to 10(coupling, hydrogenation and hydrolysis), the intermediates of Example133 were used to synthesize the following compounds of the presentinvention (as their trifluoroacetic acid salts): ##STR206##

[3-methylsulfonylamino-4-(3-phenylpropyl)-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal (Compound 134A);##STR207##

[3-methylsulfonylamino-4-(2-phenylethyl)-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 134B); ##STR208##

[3-(2,2,2-trifluoroethyl)sulfonylamino-4-(2-phenylethyl)-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 134C); ##STR209##

[3-phenylsulfonylamino-4-(2-phenylethyl)-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 134D); and ##STR210##

[3-methylaminosulfonylamino-4-(2-phenylethyl)-2-oxo-1,2-dihydro-pyridyl]acetyl-L-argininal(Compound 134E).

Example 135

Preparation of 2-Fluorobenzyl thiouronium hydrochloride ##STR211##

2-Fluorobenzyl chloride (125 g, 0.86 mole, Aldrich), thiourea (66 g,0.87 mole) and 200 mL of methanol were refluxed for 2 hours undernitrogen. The reaction was cooled and the volume was reduced to ˜40 mL.The slurry was poured into 1 liter of diethyl ether. The resulting whitesolid was filtered and dried under vacuum to give 184.5 g (97%) of thetitle compound as a white solid.

Example 136

Preparation of (2-fluorophenyl)methanesulfonyl chloride ##STR212##

The compound of Example 135 (184.5 g, 0.836 mole) was dissolved in 1700mL of distilled water. The reaction was cooled to -5° C. in an dryice/acetone bath. Chlorine gas was bubbled through the solution whilestirring with a mechanical stirrer; the reaction temperature wasmaintained at -5° C. to 5° C. throughout the addition of chlorine gas. Asodium bisulfite/water trap removed the excess chlorine gas. Thechlorine gas was added until the point of saturation was reached, therewas no rise in temperature and the reaction pale green color wasmaintained. The resulting solids were filtered and the solids weredissolved in 1 liter of ether. The ether layer was washed with dilutesodium bisulfite (NaHSO₃) four times to remove the excess chlorine. Theether layer was dried over magnesium sulfate, filtered and concentratedunder vacuum to yield 156 g (89.4%) of the title compound as a whitesolid.

Example 137

Preparation of t-Butyl (3-nitro-2-oxo-1,2-dihydropyridyl)acetate##STR213##

2-Hydroxy-3-nitro-pyridine (75 g, 0.535 mole) and 1400 mL of anhydroustetrahydrofuran were stirred at 0° C. using a mechanical stirrer.Lithium bis(trimethylsilyl)amide (1.0 M solution in tetrahydrofuran,683.5 mL) was slowly added over 30 minutes. The deep brown reactionmixture was stirred for 30 additional minutes and then t-butylbromoacetate (109.6 g, 0.561 mole) was slowly added over 30 minutes. Thereaction mixture was warmed to 25° C. overnight. The organic solventswere removed under vacuum and the residue was dissolved in 2 liters ofethyl acetate and 500 mL of water. The organic phase was dried withmagnesium sulfate, filtered and evaporated under vacuum. The residue waschromatographed on silica gel using a methylene chloride:ethanolgradient, 100:0 to 98:2, to yield 102.1 g (75%) of the title compound asa yellow-orange solid.

Example 138

Preparation of t-Butyl(3-amino-2-oxo-1,2-dihydropyridyl)acetate##STR214##

The compound of Example 137 (54 g, 0.213 mole) and 880 ml of methanoland 5 g of 10% palladium on carbon were stirred under 1 Atm of hydrogenfor 24 hours. The reaction mixture was filtered and the carbon waswashed with 200 mL of dichloromethane. The organic layer was evaporatedto yield 47.08 g (98.5%) of the title compound as a brown solid.

Example 139

Preparation oft-Butyl[3-(2-fluorobenzylsulfonyl)amino-2-oxo-1,2-dihydropyridyl]-acetate##STR215##

A mixture of the compound of Example 136 (22.27 g, 0.107 mole), thecompound of Example 138 (23.94 g, 0.107 mole), and 150 mL ofacetonitrile was cooled to 0° C. and 4-methylmorpholine (NMM) (58.68 mL,0.53 mole) was slowly added over 15 minutes. The reaction mixture waswarmed to 250° C. overnight. The solvent was evaporated under vacuum andthe residue was dissolved in 400 mL of ethyl acetate and 100 mL ofwater. The organic layer was separated and washed 3 times with 100 mL of1 N HCl, NaHCO₃ (saturated), and brine. The organic layer was dried overmagnesium sulfate, filtered and evaporated under reduced pressure. Thedark brown residue showed 3 spots by TLC. The residue was crystallizedfrom dichloromethane. Two crops of white solid were obtained, 13.70 and8.39 g, respectively. The mother liquors were evaporated and theresulting dark brown solid was crystallized from dichloromethane toyield 5.28 g of a white solid. The three crops were combined and yielded27.37 g (64.7%) of the title compound.

Example 140

Preparation of3-(2-Fluorobenzylsulfonyl)amino-2-oxo-1,2-dihydropyridyl)acetic acid##STR216##

The compound of Example 139 (13.97 g, 0.35 mole) and 50 mL ofdichloromethane were cooled to 0° C. Trifluoroacetic acid (TFA)(50 mL)was added and the reaction was stirred for 2 hours. The reaction wasjudged complete by TLC and the solvent was removed under vacuum. Toremove all traces of TFA, 100 mL of toluene and 500 mL ofdichloromethane were added and the solvents were removed under vacuum.The residue was dried under vacuum overnight to yield 11.9 g (99%) ofthe title compound.

Example 141

Preparation of[3-(2-Fluorobenzylsulfonyl)amino-2-oxo-1,2-dihydropyridyl]acetyl-N^(g)-nitro-L-argininal ethyl cyclol ##STR217##

The compound of Example 140 (22.0 g, 0.65 mole), EDC (14.87 g, 0.078mole), HOBT (10.48 g, 0.078 mole) and 250 mL of acetonitrile werestirred for 15 minutes at 25° C. This mixture was cooled to 0° C. andthe compound of Example 4, N^(g) -nitro-L-argininal ethyl cyclol (17.31g, 0.065 mol) was added. To this suspension was added slowly NMM (35.5mL, 0.323 mol). After the addition of the NMM, the reaction mixturebecame a golden brown. The reaction mixture was allowed to warm to 250°C. overnight. The solvent was removed under vacuum; the residue wasdissolved in 500 mL of dichloromethane and 100 mL of water. The organiclayer was separated and washed 3 times with 100 mL of 1 N HCl, NaHCO₃(saturated) and 100 mL of brine. The organic phase was dried overmagnesium sulfate, filtered and evaporated under reduced pressure togive 26 g of a brown foam. The residue was chromatographed on silica geleluting with dichloromethane:methanol gradient 100:0 to 97:3, to yield19.69 g (55%) of the title compound.

Example 142

Preparation of[3-(2-Fluorobenzylsulfonyl)amino-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalethyl cyclol, acetate salt ##STR218##

The compound of Example 141 (13.3 g, 0.024 mole), 80 ml ofethanol:acetic acid (4:1), and 2 g of 10% palladium on carbon werestirred overnight under 1 Atm of hydrogen. The carbon was filtered,washed with 100 mL of dichloromethane and the organic solvents wereremoved under vacuum. The resulting brown oil was dissolved in 100 mL ofdichloromethane and the solvent was removed under vacuum. The resultingglass was dried under vacuum overnight, to yield 13.6 g (>100%,theoretical yield 12.3 g) of the title compound. The product containedacetic acid trapped in the glass.

Example 143

Preparation of[3-(2-fluorobenzylsulfonyl)amino-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal##STR219##

The compound of Example 142 (13.6 g, 12.3 g theoretical yield, 0.024mole) was cooled to 0° C. and 123 mL of 8 N HCl at 0° C. was added. Thereaction mixture was stirred for 40 minutes and checked by HPLC forcompletion of the reaction. After the reaction was judged complete byHPLC, 133.9 g of sodium acetate in 150 mL of water was added to give apH of ˜4. The solution was filtered through a 0.2 micron nylon filterand chromatographed on a 4" Vydac C18 column using the following solventgradient of CH₃ CN(B) -99.9% H₂ O/0.1% TFA(A) for elution: 0% to 17% Bover 10 minutes and 17% to 23% B over 30 minutes. Two major fractions ofthe title compound were obtained: 3.86 g (33.2%)>95% pure and 4.75 g(40.8%) ˜90 to 95% pure.

Example 144

Preparation of 2-methyl-4-fluorobenzoic acid ##STR220##

To a flask fit with a reflux condenser and addition funnel was addedmagnesium (2.6 g, 106 mmol) and tetrahydrofuran (10 mL). A solution of2-bromo-5-fluorotoluene (1.0 g, 5.3 mmol) in tetrahydrofuran (2 mL) anddibromoethane (0.04 mL) was added. The solution was warmed gently in a55° C. oil bath until a vigorous reflux began. A solution of2-bromo-5-fluorotoluene (9.0 g, 48 mmol) in tetrahydrofuran (18 mL) wasadded dropwise to maintain a gentle reflux. After addition was complete,the reaction mixture was heated in a 55° C. oil bath for 1 h. Thereaction mixture was allowed to cool to room temperature and poured ontocrushed dry ice (50 mL). The excess dry ice was allowed to sublime. Thesolution was partitioned between ethyl acetate (100 mL) and 3% HCl (50mL). The organic layer was washed with brine, dried over magnesiumsulfate, and the solvent was removed in vacuo. to afford the titlecompound (8.3 g) in quantitative yield. Rf=0.34 (silica gel, 5%isopropanol/dichloromethane).

Example 145

Preparation of methyl 2-methyl-4-fluorobenzoate ##STR221##

A solution of the compound of Example 144 (8.93 g, 57.9 mmol) andsulfuric acid (0.75 mL) in methanol (150 mL) was refluxed 40 h. Thereaction mixture was cooled in an ice bath, and the acid was neutralizedto pH 7 with sodium bicarbonate. The solution was concentrated underreduced pressure. Diethyl ether and brine were added to the cruderesidue. The organic layer was dried over magnesium sulfate, and thesolvent was removed in vacuo to yield 5.1 g of the title compound (52%).Rf=0.72 (silica gel, 5% isopropanol/dichloromethane).

Example 146

Preparation of 2-carbomethoxy-5-fluorobenzyl bromide ##STR222##

A suspension of the compound of Example 145 (5.1 g, 30 mmol),N-bromosuccinimide (5.4 g, 30 mmol) and benzoyl peroxide (0.073 g, 0.30mmol) in carbon tetrachloride (115 mL) was refluxed for 18 h. Thereaction mixture was cooled to 0° C., filtered, and the precipitate waswashed with diethyl ether. The solvent was removed in vacuo to afford7.02 g (94% crude yield) of the title compound, contaminated with asmall amount of succinimide. R_(f) =0.48 20% ethyl acetate/hexanes.

Example 147

Preparation of 2-carbomethoxy-5-fluorobenzyl thiouronium hydrochloride##STR223##

The compound of Example 146 (7.016 g, 25.6 mmole based on 90% purity)and thiourea (1.9 g, 26 mmol) were refluxed in methanol (50 mL) for 48h. The residue was dissolved in water (300 mL), and washed with diethylether (3×400 mL). The solvent was removed from the aqueous layer. Theresultant yellow solid was dissolved in water (500 mL), Bio-Rex 5 resin(25 g of prewashed resin, chloride form, 8.8 meq/dry g) was added, andthe heterogenous solution was stirred for 1 h. The solution wasfiltered. The treatment with resin was repeated, and the solution wasfiltered. The solvent was removed. The residue was dissolved inmethanol, and the solvent was removed in vacuo to afford 5.65 g of alight yellow solid (79% yield).

Example 148

Preparation of 2-carbomethoxy-5-fluorobenzyl sulfonyl chloride##STR224##

The title compound was prepared using the compound of Example 147 in theprocedure of Example 136.

Example 149

Preparation of 2-carbomethoxy-6-fluorobenzyl sulfonyl chloride##STR225##

The title compound was prepared following the procedures of Examples 144through 148, except 2-methyl-5-fluorobenzoic acid was used as thestarting material in Example 144.

Example 150

Preparation of 5-2-trifluoromethyl benzenesulfonoamino-2-methyl-6-oxo-1,6-dihydro-1-pyrimidinyl)acetyl-L-argininal##STR226##

The title compound is prepared according to the procedure described inExample 113, using the intermediate resulting from the procedures inExamples 94 through 96, with the proviso that the benzylsulfonylchloride used in Example 95 is substituted with 2-trifluoromethylbenzene sulfonyl chloride (1.10 g) and 0.86 g of the compound resultingfrom Example 95 is used as starting material in Example 96.

Example 151

Preparation of 5-3-trifluoromethyl benzenesulfonoamino-2-methyl-6-oxo-1,6-dihydro-1-pyrimidinyl)acetyl-L-argininal##STR227##

The title compound is prepared according to the procedure described inExample 113, using the intermediate resulting from the procedures inExamples 94 through 96, with the proviso that the benzylsulfonylchloride used in Example 95 is substituted with 3-trifluoromethylbenzene sulfonyl chloride (1.10 g) and 0.86 g of the compound resultingfrom Example 95 is used as starting material in Example 96.

Example A

Kinetic analysis of[3[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal inan in vitro thrombin inhibition assay

The ability of a compound of a present invention,[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal(Example 10), to act as an inhibitor of thrombin catalytic activity wasassessed by determining its inhibition constant, Ki.

Enzyme activity was determined using the chromogenic substratePefachrome t-PA (CH₃ SO₂-D-hexahydrotyrosine-glycyl-L-Arginine-p-nitroaniline), obtained fromPentapharm Ltd. The substrate was reconstituted in deionized water priorto use. Purified human alpha-thrombin (3000 U/mg specific activity) wasobtained from Enzyme Research Laboratories, Inc. The buffer used for allassays was HBSA (10 mM HEPES, pH 7.5, 150 mM sodium chloride, 0.1%bovine serum albumin).

The assay for Ki determinations was conducted by combining inappropriate wells of a Corning microtiter plate, 50 microliters of HBSA,50 microliters of the test compound at a specified concentration dilutedin HBSA (or HBSA alone for V_(o) (uninhibited velocity) measurement),and 50 microliters of the chromogenic substrate (250 micromolar, 5-timesKm). At time zero, 50 microliters of alpha-thrombin diluted in HBSA wereadded to the wells, yielding a final concentration of 0.5 nM in a totalvolume of 200 microliters. Velocities of chromogenic substratehydrolysis which occurred over 60 minutes were measured by the change inabsorbance at 405 nm using a Thermo Max® Kinetic Microplate Reader. Kivalues were determined for test compounds using the relationshipsdeveloped by Williams and Morrison, Methods in Enzymology, 63:437 (1979)using steady state velocities (Vs) measured over 60 minutes. The extentof substrate hydrolysis was less than 5% over the course of this assay.

Table 1 below gives the Ki values for[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-arginial.The data show the utility of this compound as a potent in vitroinhibitor of human alpha-thrombin.

                  TABLE 1                                                         ______________________________________                                        Inhibitor constant of [3-[(benzylsulfonyl)                                      amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-                                     argininal against human alpha-thrombin                                        amidolytic activity                                                              Compound                Ki (pM)                                          ______________________________________                                        [3-[(benzylsulfonyl)amino]-2-oxo-1,2-                                                                  289 ± 32                                            dihydropyridyl]acetyl-L-argininal                                           ______________________________________                                    

Example B

In vitro Enzyeme Assays for Speciticity Determination

The ability of compounds of the present invention to act as a selectiveinhibitor of thrombin catalytic activity was assessed by determining theconcentration of compound which inhibited the actvity of this enzyme by50%, (IC₅₀), and comprising this value to that determined for all orsome of the following related serine proteases: recombinant tissueplasminogen activator (rt-PA), plasmin, activated protein C,chymotrypsin, factor Xa and trypsin.

The buffer used for all assays was HBSA (10 mM HEPES, pH 7.5, 150 mMsodium chloride, 0.1% bovine serum albumin).

The assay for IC₅₀ determinations was conducted by combining inappropriate wells of a Corning microtiter plate, 50 microliters of HBSA,50 microliters of the test compound at a specified concentration(covering a broad concentration range) diluted in HBSA (or HBSA alonefor V_(O) (uninhibited velocity) measurement), and 50 microliters of theenzyme diluted in HBSA. Following a 30 minute incubation at ambienttemperature, 50 microliters of the substrate at the concentrationsspecified below were added to the wells, yielding a final total volumeof 200 microliters. The initial velocity of chromogenic substratehydrolysis was measured by the change in absorbance at 405 nm using aThermo Max® Kinetic Microplate Reader over a 5 minute period in whichless than 5% of the added substrate was utilized. The concentration ofadded inhibitor which caused a 50% decrease in the initial rate ofhydrolysis was defined as the IC₅₀ value.

Thrombin (fIIa) Assay

Enzyme activity was determined using the chromogenic substrate,Pefachrome t-PA (CH₃ SO₂-D-hexahydrotyrosine-glycyl-L-Arginine-p-nitroaniline, obtained fromPentapharm Ltd.). The substrate was reconstituted in deionized waterprior to use. Purified human a-thrombin was obtained from EnzymeResearch Laboratories, Inc. The buffer used for all assays was HBSA (10mM HEPES, pH 7.5, 150 mM sodium chloride, 0.1% bovine serum albumin).

IC₅₀ determinations were conducted where HBSA (50 mL), a-thrombin (50μl) and inhibitor (50 μl) (covering a broad concentration range), werecombined in appropriate wells and incubated for 30 minutes at roomtemperature prior to the addition of substrate Pefachrome-t-PA (50 μl).The initial velocity of Pefachrome t-PA hydrolysis was measured by thechange in absorbance at 405 nm using a Thermo Max® Kinetic MicroplateReader over a 5 minute period in which less than 5% of the addedsubstrate was utilized. The concentration of added inhibitor whichcaused a 50% decrease in the initial rate of hydrolysis was defined asthe IC₅₀ value.

Factor Xa

Factor Xa catalytic activity was determined using the chromogenicsubstrate S-2765(N-benzyloxycarbonyl-D-arginine-L-glycine-L-arginine-p-nitroaniline),obtained from Pharmacia Upjohn (Franklin, Ohio). All substrates werereconstituted in deionized water prior to use. The final concentrationof S-2765 was 250 μM (about 5-times Km). Purified human Factor X wasobtained from Enzyme Research Laboratories, Inc. (South Bend, Ind.) andFactor Xa (FXa) was activated and prepared from it as described [Bock,P. E., Craig, P. A., Olson, S. T., and Singh, P. Arch. Biochem. Biophys.273:375-388 (1989)].

Recombinant tissue plasminogen activator (rt-PA) Assay

rt-PA catalytic activity was determined using the substrate, Pefachromet-PA (CH₃ SO₂ -D-hexahydrotyrosine-glycyl-L-arginine-p-nitroaniline,obtained from Pentapharm Ltd.). The substrate was made up in deionizedwater followed by dilution in HBSA prior to the assay in which the finalconcentration was 500 micromolar (about 3-times Km). Human rt-PA(Activase®) was obtained from Genentech Inc. The enzyme wasreconstituted in deionized water and diluted into HBSA prior to theassay in which the final concentration was 1.0 nM.

Plasmin Assay

Plasmin catalytic activity was determined using the chromogenicsubstrate, S-2366 [L-pyroglutomyl-L-prolyl-L-arginine-p-nitroanilinehydrochloride], which was obtained from Pharmacia-Upjohn. The substratewas made up in deionized water followed by dilution in HBSA prior to theassay in which the final concentration was 300 micromolar (about2.5-times Km). Purified human plasmin was obtained from Enzyme ResearchLaboratories, Inc. The enzyme was diluted into HBSA prior to assay inwhich the final concentration was 1.0 nM.

Activated Protein C (aPC) Assay

aPC catalytic activity was determined using the chromogenic substrate,Pefachrome PC(delta-carbobenzloxy-D-lysine-L-prolyl-L-arginine-p-nitroanilinedihydrochloride), obtained from Pentapharm Ltd.). The substrate was madeup in deionized water followed by dilution in HBSA prior to the assay inwhich the final concentration was 250 micromolar (about 3-times Km).Purified human aPC was obtained from Hematologic Technologies, Inc. Theenzyme was diluted into HBSA prior to assay in which the finalconcentration was 1.0 nM.

Chymotrypsin Assay

Chymotrypsin catalytic activity was determined using the chromogenicsubstrate, S-2586(methoxy-succinyl-L-arginine-L-prolyl-L-tyrosyl-p-nitroanilide), whichwas obtained from Pharmacia-Upjohn. The substrate was made up indeionized water followed by dilution in HBSA prior to the assay in whichthe final concentration was 100 micromolar (about 9-times Km). Purified(3X-crystallized;CDI) bovine pancreatic alpha-chymotrypsin was obtainedfrom Worthington Biochemical Corp. The enzyme was reconstituted indeionized water and diluted into HBSA prior to assay in which the finalconcentration was 1.0 nM.

Trypsin Assay

Trypsin catalytic activity was determined using the chromogenicsubstrate, S-2222 (benzoyl-L-isoleucine-L-glutamic acid-[gamma-methylester]-L-arginine-p-nitroanilide), which was obtained fromPharmacia-Upjohn. The substrate was made up in deionized water followedby dilution in HBSA prior to the assay in which the final concentrationwas 250 micromolar (about 4-times Km). Purified (3X-crystallized; TRL3)bovine pancreatic trypsin was obtained from Worthington BiochemicalCorp. The enzyme was reconstituted in deionized water and diluted intoHBSA prior to assay in which the final concentration was 0.5 nM.

Tables 2, 3A and 3B list the determined IC₅₀ values for certain of theenzymes listed above and demonstrate the high degree of specificity forthe inhibition of alpha-thrombin compared to these related serineproteases.

                  TABLE 2                                                         ______________________________________                                        IC.sub.50 values (nM) for the inhibition of                                     human alpha thrombin amidolytic activity compared to selected                 serine proteases for compounds of Example 10 (column A),                      Example 90, compound B (column B), and Example 113,                           compounds C, D, and E (columns C, D, and E, respectively)                               Enzyme                                                                            A        B    C      D    E                                   ______________________________________                                        Alpha-thrombin                                                                            0.66     0.98   0.467  2.32 141                                     rt-PA NI* NI* ND   NI* NI*                                                    Plasmin NI* NI* NI* NI* NI*                                                   aPC NI* NI* ND  NI* NI*                                                     ______________________________________                                         NI*--IC.sub.50 value > 2500 nM.                                               ND--not determined                                                       

                  TABLES 3A and 3B                                                ______________________________________                                        IC.sub.50 values (nM) for inhibition of human alpha thrombin amidolytic        activity compared to inhibition of rt-PA, plasmin, and aPC for compounds      made according to Examples 89 and 90 with the stated R.sub.1 substitution    ______________________________________                                        Table 3A                                                                                      Thrombin rt-PA   Plasmin                                                                             aPC                                      R.sub.1 substitution (IC.sub.50) (IC.sub.50) (IC.sub.50) (IC.sub.50)        ______________________________________                                          2-CF.sub.3 -phenyl 5.6  NI* NI* NI*                                           3-CF.sub.3 -phenyl 3.1  NI* NI* NI*                                           2-Me-phenyl 1.4  NI* NI* NI*                                                  3-Me-phenyl 0.85 NI* NI* NI*                                                  2-Me,5-F-phenyl 1.97 NI* NI* NI*                                              2-OMe-phenyl 1.98 NI* NI* NI*                                                 3-OMe-phenyl 0.65 NI* NI* NI*                                                 2-CMe,5-Cl-phenyl 1.16 NI* NI* NI*                                            2-NH.sub.2 -phenyl 3.7  NI* NI* NI*                                         ______________________________________                                        Table 3B                                                                        Compound of Example                                                                         Thrombin rtPA    Plasmin                                                                             aPC                                    ______________________________________                                          121A 14.7 --  >2500 --                                                        121E .597 inactive >2500 inactive                                             121G 46.9 -- ˜2500 --                                                   121J and 143 .763 >2500 >2500 inactive                                        121K .882 inactive >2500 inactive                                             121M .623 -- >2500 --                                                         121P 1.73 >2500 >2500 inactive                                                126B .882 inactive >2500 inactive                                             126C .519 inactive >2500 inactive                                             126F .71 inactive ˜2500 inactive                                        134B 7.95 >2500 >2500 inactive                                              ______________________________________                                    

Example C

Ex vivo anticoagulant effects of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalin human plasma

The ex vivo anticoagulant effect of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalwas determined by measuring the prolongation of the activated partialthromboplastin time (APTT) over a broad concentration range of the addedinhibitor, using pooled normal human plasma. Fresh frozen citratedpooled normal human plasma was obtained from George King Biomedical,Overland Park, Kans. Measurements of APTT were made using theCoag-A-Mate RA4 automated coagulometer (General Diagnostics, OrganonTechnica, Oklahoma City, Okla.) using the Platelin® L reagent (OrganonTechnica, Durham, N.C.) as the initiator of clotting according to themanufacturer's instructions. The assay was conducted by making a seriesof dilutions of the test compounds in rapidly thawed plasma followed byadding 200 microliters to the wells of the assay carousel. As shown inFIG. 6, a compound of the present invention,[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,prolonged the APTT in a dose dependent manner in human plasmademonstrating an anticoagulant effect in this species of mammals.

Example D

Evaluation of the antithrombotic potential of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalin an experimental rat model of thrombosis

The demonstrated anticoagulant effects of a compound of the presentinvention,[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,in human citrated plasma indicated that this compound may have potentantithrombotic effects in an experimental model of thrombosis. Toinvestigate this, the antithrombotic (prevention of thrombus formation)properties of this compound were evaluated using the followingestablished experimental model of acute vascular thrombosis.

Rat model of FeCl₃ -induced platelet-dependent arterial thrombosis

This is a well characterized model of platelet dependent, arterialthrombosis which has been used to evaluate the potential ofantithrombotic compounds such as direct thrombin inhibitors. Kurz, K.D., Main, B. W., and Sandusky, G. E., Thromb. Res., 60:269-280 (1990).In this model a platelet-rich, occlusive thrombus is formed in a segmentof the rat carotid artery treated locally with a fresh solution of FeCl₃absorbed to a piece of filter paper. The FeCl₃ is thought to diffuseinto the treated segment of artery and cause de-endothelialization ofthe affected vessel surface. This results in the exposure of blood tosubendothelial structures which in turn causes platelet adherence,thrombin formation and platelet aggregation resulting in occlusivethrombus formation. The effect of a test compound on the incidence ofocclusive thrombus formation following the application of the FeCl₃ ismonitored by ultrasonic flowtometry and is used as the primary endpoint. The use of flowtometry to measure carotid artery blood flow, is amodification of the original procedure in which thermal detection ofclot formation was employed. Kurz, K. D., Main, B. W., and Sandusky, G.E., Thromb. Res., 60:269-280 (1990).

Male Harlan Sprague Dawley rats (420-450 g) were acclimated at least 72hours prior to use and fasted for 12 hours prior to surgery with freeaccess to water. The animals were prepared, anesthetized with Nembutalfollowed by the insertion of catheters for blood pressure monitoring,drug and anesthesia delivery. The left carotid artery was isolated bymaking a midline cervical incision followed by blunt dissection andspreading techniques to separate a 2 cm segment of the vessel from thecarotid sheath. A silk suture is inserted under the proximal and distalends of the isolated vessel to provide clearance for the placement of aultrasonic flow probe (Transonic) around the proximal end of the vessel.The probe is then secured with a stationary arm.

Following surgery the animals were randomized in either a control(saline) or treatment group with test compound,[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,with at least 6 animals per group per dose. The test compound wasadministered as a single intravenous bolus at the doses outlined inTable 4 after placement of the flow probe and 5 minutes prior to thethrombogenic stimulus. At t=0, a 3 mm diameter piece of filter paper(Whatman #3) soaked with 10 microliters of a 35% solution of fresh FeCl₃(made up in water) was applied the segment of isolated carotid arterydistal to the flow probe. Blood pressure, blood flow, heart rate, andrespiration were monitored for 60 minutes.

The incidence of occlusion (defined as the attainment of zero bloodflow) was recorded as the primary end point.

The efficacy of the[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalas an antithrombotic agent in preventing thrombus formation in this invivo model was demonstrated by the reduction in the incidence ofthrombotic occlusion as shown in Table 4 below.

                  TABLE 4                                                         ______________________________________                                        Results of [3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydro-                        pyridyl]actyl-L-argininal in the FeCl.sub.3 Model of Thrombosis in Rats                         Dose          Incidence                                                                      Treatment Group (mg/kg) n of Occulusion    ______________________________________                                          Saline -- 6 6/6                                                             [3-[(benzylsulfonyl)amino]-                                                                   0.3       6     6/6                                             2-oxo-1,2-                                                                    dihydropyridyl]acetyl-L-                                                      argininal                                                                     [3-[(benzylsulfonyl)amino]- 1.0 6 3/6                                         2-oxo-1,2-                                                                    dihydropyridyl]acetyl-L-                                                      argininal                                                                     [3-[(benzylsulfonyl)amino]- 3.0 6  1/6*                                       2-oxo-1,2-                                                                    dihydropyridyl]acetyl-L-                                                      argininal                                                                     [3-[(benzylsulfonyl)amino]- 5.0 6  0/6*                                       2-oxo-1,2-                                                                    dihydropyridyl]acetyl-L-                                                      argininal                                                                   ______________________________________                                         *p ≦ 0.05 from saline control by Fishers test                     

The effective dose which prevents 50% of thrombotic occlusions in thismodel (ED₅₀) can be determined from the above data by plotting theincidence of occlusion versus the dose administered. This allows adirect comparison of the antithrombotic efficacy of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,with other antithrombotic agents which have also been evaluated in thismodel as described above. Table 5 lists the ED₅₀ values for several wellknown anticoagulant agents in this model compared to[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal.

                  TABLE 5                                                         ______________________________________                                        Efficacy of [3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]-              acetyl-L-argininal compared to other antithrombotic agents based on          ED.sub.50                                                                       for thrombus prevention in the FeCl.sub.3 model of arterial thrombosis      in rats                                                                           Compound                ED.sub.50.sup.a                                   ______________________________________                                        Standard Heparin        200    U/kg                                             Argatroban 3.8 mg/kg                                                          Hirulog ™ 3.0 mg/kg                                                        [3-[(benzylsulfonyl)amino]-2-oxo-1,2- 1.0 mg/kg                               dihydropyridyl]acetyl-L-argininal                                           ______________________________________                                         .sup.a ED.sub.50 is defined as the dose that prevents the incidence of        complete thrombotic occlusion in 50% of animals tested.                  

The data presented in Table 4 clearly demonstrate the effectiveness of acompound of the present invention,[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,in preventing occlusive thrombus formation in this experimental model.The relevance of these data to preventing human thrombosis can beinferred from the comparison to the other anticoagulant agents listed inTable 5 which have been evaluated in an identical manner in thisexperimental model and have demonstrated antithrombotic efficacy inpreventing thrombus formation clinically as described in the followingliterature citations: Heparin-Hirsh, J., N. Engl. J. Med., 324:1565-1574(1992) and Cairns, J. A. et al., Chest, 102:456S-481S (1992);Argatroban-Gold, H. K. et al., J. Am. Coll. Cardiol., 21:1039-1047(1993); and Hirulog™-Sharma, G. V. R. K. et al., Am. J. Cardiol.,72:1357-1360 (1993) and Lidon, R. M. et al., Circulation, 88:1495-1501(1993). The in vivo comparison of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalwith the clinically effective antithrombotic agents, Standard Heparin,Argatroban, and Hirulog™, in the same rodent model of experimentalthrombosis, coupled with the demonstrated anticoagulant effects of[3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininalin human plasma described above in Example C would lead one skilled inthe art to conclude that the compounds of the present invention will bean effective antithrombotic agent in humans.

Example E

Multiple Extracorporeal Shunt Model in Rats Utilizing Oral Dosing

The compounds of Example 10 and 143 were evaluated in a multichamber A-Vshunt model in rats. The A-V shunt model is one of the most common andgenerally used systems to evaluate antithrombotic compounds. Smith, J.R. and White, A. M. Br. J. Pharmacol., 77:29-38 (1982). In this model alocalized clot made up of primarily fibrin with some platelet andmacrophage involvement (Shand, R. A. and Smith, J. R. and Wallis, R. B.Thromb. Res., 36:223-232 (1984)), is formed on an artificialthrombogenic surface (typically a segment of silk or cotton thread)contained in a sialstic chamber which is part of an exteriorized shuntbetween the carotid artery and jugular vein. The procedure described inthis Example is a modified A-V shunt model that allows for oral dosingof test agents and subsequent evaluation of efficacy over a two to threehour window in time.

Briefly, male Harlan Sprague Dawley rats (420-450 g) were acclimated atleast 72 hours prior to use. The animals were fasted for 12 hours priorto surgery with free access to water. Unanesthetized animals weregrouped into three or four dosage groups (six or seven animals pergroup) and administered test agents orally via gavage needle, at dosesof 1.0, 3.0, 10 and 50 mg/kg for the compound of Example 10, and 3.0, 10and 30 mg/kg for the Compound of Example 143. Immediately after oraldosing, animals were anesthetized with sodium pentobarbital (Nembutal)given intraperitoneally at a dose of 50 mg/kg body weight, and placed ona isothermal pad to maintain body temperature. The level of anesthesiawas monitored every 15 minutes by neuro-response to a tail pinch,respiration and core temperature. The desired depth of surgicalanesthesia was maintained by administering subsequent doses (5 mg/kg)intravenously. The left femoral artery was catheterized using standardprocedures for blood pressure monitoring and blood sampling, withpolyethylene tubing (PE50). The left femoral vein was catheterized withPE50 tubing for delivery of anethestic.

The exteriorized shunts were assembled by connecting two pieces ofsaline filled 12.5 cm PE90 tubing with a 6 cm piece of PE160 tubingcontaining a 6 cm piece of silk suture size 3 and clamped withhemostats. A small 0.5 cm portion of the silk thread protrudes from thejunction of the chamber with the shunt. The left jugular vein and rightcarotid artery were catheterized with the ends of the PE90 shunt. Theshunt was unclamped and blood allowed to flow from the carotid artery,through the chamber, and exit the shunt via the jugular vein. After 15minutes, both sides of the chamber were clamped and the suturecontaining the clot removed following detachment of the arterial end ofthe chamber. The clot was immediately weighed and recorded. Thisprocedure takes place at predetermined intervals (60, 90, 120, and 150minutes after oral dosing) to allow assessment of efficacy over a largewindow in time. Four shunts were placed with flow initiated at 45, 75,105, and 135 minutes after oral compound administration. Clot weightfrom the four shunts was the primary endpoint of the protocol. Bloodpressure, heart rate core temperature and respiration were monitoredcontinuously. Following termination of the experiment the animals wereeuthanized with a 120 mg/kg dose of Nembutal. One experiment wasperformed per animal.

ED₅₀ values were calculated at 60, 90, 120, and 150 minutes after oraldosing of test compound. ED₅₀ is that dose that reduced the clot size by50%. For the compound of Examples 10 and 143, the ED₅₀ values were asshown in Table 6, below, and demonstrate the oral availability andefficacy of the compounds.

                  TABLE 6                                                         ______________________________________                                        Time after oral dose                                                                         ED.sub.50                                                      ______________________________________                                         60 min        (Example 10)                                                                             (Example 143)                                          <1.0 mg/kg   22 mg/kg                                                         90 min 2.9 mg/kg 20 mg/kg                                                    120 min 2.9 mg/kg 24 mg/kg                                                    150 min 8.2 mg/kg 28 mg/kg                                                  ______________________________________                                    

We claim:
 1. A compound of formula: ##STR228## wherein (a) X is selectedfrom the group consisting of --S(O)₂ --, --N(R')--S(O)₂ --, --(C═O)--,--OC(═O)--, --NH--C(═O)--, --P(O) (R")-- and a direct link, wherein R'is hydrogen, alkyl of 1 to about 4 carbon atoms, aryl of about 6 toabout 14 carbon atoms or aralkyl of about 6 to about 16 carbon atoms,and R" is NR', OR', R', or SR', with the proviso that R" is not NH, OH,H, or SH, and;(b) R₁ is selected from the group consisting of:(1) alkylof 1 to about 12 carbon atoms, (2) alkyl of 1 to about 3 carbon atomssubstituted with cyclic alkyl of about 3 to about 8 carbon atoms, whichoptionally is substituted in the ring carbons with hydroxyl, amino,guanidino, amidino, or alkoxyl or alkyl each of 1 to about 3 carbons,(3) cyclic alkyl of 3 to about 15 carbon atoms, which optionally issubstituted in the ring carbons with hydroxyl, amino, guanidino,amidino, or alkoxyl or alkyl each of 1 to about 3 carbons, (4)heterocycloalkyl of 4 to about 10 ring atoms with the ring atomsselected from carbon and heteroatoms, wherein the heteroatoms areselected from the group consisting of oxygen, nitrogen, and S(O)_(i),wherein i is 0, 1 or 2, and which is optionally substituted on the ringcarbons with hydroxyl, alkoxyl or alkyl of 1 to about 3 carbons, amino,guanidino, or amidino, (5) heterocyclo of 4 to about 10 ring atoms withthe ring atoms selected from carbon and heteroatoms, wherein theheteroatoms are selected from the group consisting of oxygen, nitrogen,and S(O)i, wherein i is 0, 1 or 2, and including ##STR229## wherein##STR230## is a 5 to 7 member heterocycle of 3 to 6 ring carbon atoms,where V is --CH₂ --, --O--, --S(═O)--, --S(O)₂ -- or --S--,optionallysubstituted on the ring carbons with hydroxyl, alkoxyl or alkyl each of1 to about 3 carbons, amino, guanidino, or amidino, (6) alkenyl of 2 toabout 6 carbon atoms which is optionally substituted with cyclic alkylof about 3 to about 8 carbon atoms, which optionally is substituted inthe ring carbons with hydroxyl, amino, guanidino, amidino, or alkoxyl oralkyl each of 1 to about 3 carbons, (7) aryl of about 6 to about 14carbon atoms which is optionally mono-, di- or tri-substituted with Y₁,Y₂, and/or Y₃, (8) heteroaryl of 5 to 14 atoms with the ring atomsselected from carbon and heteroatoms, wherein the heteroatoms areselected from oxygen, nitrogen, and S(O)_(i), wherein i is 0, 1 or 2,and which is optionally mono-, di- or tri-substituted with Y₁, Y₂,and/or Y₃, (9) aralkyl of about 7 to about 15 carbon atoms which isoptionally substituted on the alkyl chain with hydroxy or halogen andoptionally mono-, di-, or tri-substituted on the aryl ring with Y₁, Y₂,and/or Y₃, (10) heteroaralkyl of 6 to 11 atoms with the ring atomsselected from carbon and heteroatoms, wherein the heteroatoms areselected from oxygen, nitrogen, and S(O)_(i), wherein i is 0, 1 or 2,and which is optionally substituted on the alkyl chain with hydroxy orhalogen and optionally mono-, di- or tri-substituted on the ring withY₁, Y₂, and/or Y₃, (11) aralkenyl of about 8 to about 16 carbon atomswhich is optionally mono-, di-, or tri-substituted on the aryl ring withY₁, Y₂, and/or Y₃, (12) heteroaralkenyl of 7 to 12 atoms with the ringatoms selected from carbon and heteroatoms, wherein the heteroatoms areselected from oxygen, nitrogen, and S(O)_(i), wherein i is 0, 1 or 2,and which is optionally mono-, di- or tri-substituted on the ring withY₁, Y₂, and/or Y₃, ##STR231## (17) difluoromethyl and perfluoroalkyl of1 to about 12 carbon atoms, (18) perfluoroaryl of about 6 to about 14carbon atoms, (19) perfluoroaralkyl of about 7 to about 15 carbon atoms,and (20) hydrogen,wherein Y₁, Y₂, and Y₃ are (i) independently selectedfrom the group consisting of hydrogen, halogen, cyano, tetrazolyl,amino, guanidino, amidino, methylamino, and methylguanidino, --CF₃,--CF₂ H, --CF₂ CF₃, --CH(CF₃)₂, --C(OH)(CF₃)₂, --OCF₃, OCF₂ CF₃,--OC(O)NH₂, --OC(O)NHZ₁, --OC(O)NZ₁ Z₂, --NHC(O)Z₁, --NHC(O)NH₂,--NHC(O)NZ₁, --NHC(O)NZ₁ Z₂, --C(O)OH, --C(O)NH₂, --C(O)NHZ₁, --C(O)OZ₁,--P(O)₃ H, --P(O)₃ H₂, --P(O)₃ (Z₁)₂, --S(O)₃ H , --S(O)_(m) Z₁, --Z₁,--OZ₁, --OH, --NH₂, --NHZ₁, and --NZ₁ Z₂, wherein m is 0, 1 or 2, and Z₁and Z₂ are independently selected from the group consisting of alkyl of1 to about 12 carbon atoms, aryl of about 6 to about 14 carbon atoms,heteroaryl of about 5 to about 14 atoms having 1 to about 9 carbonatoms, aralkyl of about 7 to about 15 carbon atoms, and heteroaralkyl ofabout 6 to about 11 atoms having about 3 to about 9 carbon atoms, or(ii) Y₁ and Y₂ are selected together to be --OC(Z₃)(Z₄)O--, wherein Z₃and Z₄ are independently selected from the group consisting of hydrogen,alkyl of 1 to about 12 carbon atoms, aryl of about 6 to about 14 carbonatoms heteroaryl of about 5 to about 14 atoms having 1 to about 9 carbonatoms, aralkyl of about 7 to about 15 carbon atoms, and heteroaralkyl ofabout 6 to about 11 atoms having about 3 to about 9 carbon atoms, withthe proviso that if X is not a direct link, then R₁ is not hydrogen, (c)R₂ is selected from the group consisting of hydrogen, alkyl of 1 toabout 4 carbon atoms, and alkenyl of about 2 to about 4 carbon atoms,(d) R₃ is selected from the group consisting of ##STR232## where W isnitrogen or carbon; (e) Het is ##STR233## wherein (1) R₄ is selectedfrom the group consisting of(a) R₁, --OR₁, --NHR₁, --S(O)_(n) R₁, andhalogen, wherein n is 0, 1 or 2, and R₁ is independently selected, withthe proviso that R₄ is not a camphor derivative or ##STR234##heterocyclo group, (b) alkyl of 1 to about 12 carbon atoms substitutedwith Z₅ wherein Z₅ is selected from the group consisting of hydroxy,halogen, --C(O)OH, --C(O)OR₈, --S(O)₃ OH, and --S(O)_(p) R₈ wherein R₈is alkyl of 1 to about 6 carbon atoms and p is 0, 1 or 2, and (c)alkenyl of about 3 to about 6 carbon atoms; (2) R5 is selected from thegroup consisting of(a) hydrogen, (b) alkyl of 1 to about 10 carbonatoms, (c) alkyl of 1 to about 3 carbon atoms substituted with cyclicalkyl of about 3 to about 8 carbon atoms, (d) cyclic alkyl of 3 to about6 carbon atoms, (e) heterocycloalkyl of 4 to about 6 ring atoms with thering atoms selected from carbon and heteroatoms, wherein the heteroatomsare selected from the group consisting of oxygen, nitrogen and--S(O)_(i) -- wherein i is independently 0, 1 or 2, (f) heterocyclo of 4to about 6 ring atoms with the ring atoms selected from carbon atoms andheteroatoms, wherein the heteroatoms are selected from the groupconsisting of oxygen, nitrogen and --S(O)_(i) -- wherein i isindependently 0, 1 or 2 and which is attached to Het by a ring carbonatom, (g) alkenyl of 2 to about 6 carbon atoms which is optionallysubstituted with cyclic alkyl of 3 to about 5 carbon atoms, (h) arylwhich is optionally mono-, di- or tri- substituted with Y₁, Y₂ and/or Y₃respectively, (i) heteroaryl of 5 to 6 atoms with the ring atomsselected from carbon atoms and heteroatoms, wherein the heteroatoms areselected from the group consisting of oxygen, nitrogen, and --S(O)_(i)-- wherein i is independently 0, 1 or 2 and which is optionally mono-,di- or tri- substituted with Y₁, Y₂ and/or Y₃, (j) aralkyl of about 7 toabout 10 carbon atoms which is optionally mono-, di- or tri-substitutedon the aryl ring with Y₁, Y₂ and/or Y₃ ; (k) heteroaralkyl of 6 to 9atoms with the ring atoms selected from carbon atoms and heteroatoms,wherein the heteroatoms are selected from the group consisting ofoxygen, nitrogen and --S(O)_(i) -- wherein i is independently 0, 1 or 2and which is optionally mono-, di- or tri- substituted on the ring withY₁, Y₂ and/or Y₃, (l) aralkenyl of 8 carbon atoms which is optionallymono-, di- or tri- substituted on the aryl ring with Y₁, Y₂ and/or Y₃,(m) heteroaralkenyl of 7 to 8 atoms with the ring atoms selected fromcarbon atoms and heteroatoms, wherein the heteroatoms are selected fromthe group consisting of oxygen, nitrogen, and --S(O)_(i) -- wherein i isindependently 0, 1 or 2, and which is optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃, (n) halogen, (o)difluoromethyl or perfluoroalkyl of 1 to 3 carbon atoms, (p)perfluorophenyl, (q) perfluoroaralkyl of 7 to about 9 carbon atoms, and(r) alkoxy of 1 to about 10 carbon atoms; (3) R₆ is selected from thegroup consisting of(a) R₁, --OR₁, --NHR₁, --S(O)_(n) R₁, or halogen,wherein n is 0, 1 or 2, and R₁ is independently selected, with theproviso that R₆ is not a camphor derivative or ##STR235## heterocyclegroup, and (b) alkyl of 1 to about 12 carbon atoms substituted with Z₆,wherein Z₆ is selected from the group consisting of hydroxy, halogen,--OR₉, --NHR₉, --C(O)OH, --C(O)OR₉, --S(O)₂ OH and --S(O)_(p) R₉ whereinR₉ is selected from alkyl of 1 to about 12 carbon atoms, aryl of about 6to about 10 carbon atoms optionally mono-, di or tri-substituted on thering with Y₁, Y₂ and/or Y₃, aralkyl of about 7 to about 12 carbon atomsoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂ and/orY₃, heteroaryl of 1 to about 9 carbon atoms with the ring atoms selectedfrom carbon and heteroatoms selected from the group consisting ofoxygen, nitrogen and --S(O)_(p) -- and optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃ ; and heteroaralkyl ofabout 2 to about 10 carbon atoms with the ring atoms selected fromcarbon and heteroatoms selected from the group consisting of oxygen,nitrogen and --S(O)_(p) -- and optionally mono-, di- or tri-substitutedon the ring with Y₁, Y₂ and/or Y₃ ; and (4) R7 is independently selectedfrom the R5 group of substituents, provided that R7 is not halogen; andpharmaceutically acceptable salts thereof.
 2. A compound according toclaim 1, wherein X is selected from the group consisting of --SO₂ --,--NH--S(O)₂ --, and --N(R')--S(O)₂ --.
 3. A compound according to claim2, wherein X is --SO₂ --.
 4. A compound according to claim 1, wherein R₁is selected from the group consisting of alkyl, cycloalkyl, aralkyl, andaryl.
 5. A compound according to claim 4, wherein R₁ is selected fromthe group consisting of substituted or unsubstituted phenyl, benzyl, andnaphthyl.
 6. A compound according to claim 5, wherein R₁ has one or twosubstituents selected from the group consisting of methyl, methoxy,fluoro, chloro, trifluoromethyl, and --OCF₃.
 7. A compound according toclaim 4, wherein R₁ is cyclohexyl or cyclohexylmethyl.
 8. A compoundaccording to claim 1, wherein R₂ is hydrogen.
 9. A compound according toclaim 1 wherein R₃ is ##STR236##10.
 10. A compound according to claim 9,wherein R₄ is selected from the group consisting of: (a) hydrogen,(b)alkyl of 1 to 6 carbon atoms or alkyl of 1 to 6 carbon atoms substitutedwith Z₅, wherein Z₅ is selected from the group consisting of hydroxy,halogen, --C(O)OH, --C(O)OR₈, --S(O)₂ OH and --S(O)_(p) R₈, (c) alkyl of1 to 3 carbon atoms substituted with cyclic alkyl of 3 to 5 carbonatoms, (d) alkenyl of about 3 to about 6 carbon atoms, (e) cycloalkyl ofabout 3 to about 5 carbon atoms, (f) heteroaryl of 5 atoms, and (g)heteroaralkyl of 6 atoms.
 11. A compound according to claim 9, whereinR₅ is selected from the group consisting of hydrogen, alkyl of 1 toabout 5 carbon atoms, trifluoromethyl, and alkoxy of 1 to 4 carbonatoms.
 12. A compound according to claim 11, wherein R₅ is hydrogen. 13.A compound according to claim 9, wherein R₆ is selected from the groupconsisting of:(a) hydrogen, (b) alkyl of 1 to about 12 carbon atoms oralkyl of 1 to 12 carbon atoms substituted with Z₆, wherein Z₆ isselected from the group consisting of hydroxy, halogen, --OR₉, --NHR₉,--C(O)OH, --C(O)OR₉, --S(O)₂ OH and --S(O)_(p) R₉, (c) alkyl of 1 toabout 3 carbon atoms substituted with cyclic alkyl of about 5 to about 8carbon atoms, (d) alkenyl of about 2 to about 6 carbon atoms which isoptionally substituted with cyclic alkyl of about 3 to about 8 carbonatoms, or aryl of about 6 to about 10 carbon atoms, (e) aralkyl orsubstituted aralkyl, (f) heteroaralkyl of about 5 to 10 ring atoms orsubstituted heteroaralkyl of about 5 to 10 ring atoms, (g) aralkenyl ofabout 8 to 15 carbon atoms which is optionally mono-, di- ortri-substituted on the ring with Y₁, Y₂ and/or Y₃, and (h)heteroaralkenyl of about 5 to 10 ring atoms or substitutedheteroaralkenyl of about 5 to 10 ring atoms.
 14. A compound according toclaim 13, wherein R₄ and R₅ are hydrogen or methyl and R₆ is selectedfrom the group consisting of aralkyl of about 8 to about 13 carbonatoms, and --O-aralkyl, --NH-aralkyl, or --S(O)_(p) -aralkyl all ofabout 7 to about 12 carbon atoms.
 15. A compound according to claim 14,wherein the aryl portion of the aralkyl group of R₆ is selected fromunsubstituted or substituted phenyl or naphthyl.
 16. A compoundaccording to claim 15, wherein said substituents of the aryl ring areselected from the group consisting of methyl, methoxy, fluoro, chloroand trifluoromethyl.
 17. A compound according to claim 13, wherein R₆ isselected from the group consisting of phenylethyl, phenylpropyl,cyclohexylethyl and cyclohexylpropyl.
 18. A compound according to claim9, wherein R₇ is selected from the group consisting of hydrogen, methyl,difluoromethyl and trifluoromethyl.
 19. A compound according to claim18, wherein R₇ is hydrogen.
 20. A compound according to claim 9, whereinHet is ##STR237## wherein R₄ is selected from the group consisting ofhydrogen, methyl, ethyl, propenyl, allyl, propyl, isopropyl, butyl,R-sec-butyl, S-sec-butyl, isobutyl, 1-pentyl, R-2-pentyl, S-2-pentyl,3-pentyl, S-1-(2-methyl)-butyl, R-2-(3-methyl)-butyl,1-(3-methyl)-butyl, R-1-(2-methyl)-butyl, cyclopentyl, 2-pyrolyl,3-pyrolyl, 1-hexyl, S-2-hexyl, R-2-hexyl, R-3-hexyl, and S-3-hexyl; andR₅ and R₆ are independently selected from hydrogen and methyl.
 21. Acompound according to claim 20, wherein R₄ is hydrogen or methyl.
 22. Acompound according to claim 9, wherein X is --S(O)₂ --, R₁ issubstituted or unsubstituted aralkyl or substituted or unsubstitutedphenyl, and Het is ##STR238##
 23. A compound according to claim 22,wherein R₁ is substituted or unsubstituted benzyl or phenyl.
 24. Acompound according to claim 9 wherein X is --S(O)₂ --.
 25. A compoundaccording to claim 24 wherein R₁ is alkyl, aryl or aralkyl.
 26. Acompound according to claim 25 wherein R₁ is aryl or aralkyl optionallysubstituted with Y₁ and/or Y₂ and Y₁ and Y₂ are independently selectedfrom --C(O)OH, --C(O)OZ₁, --OH, --S(O)_(m) Z₁, and --CF₃.
 27. A compoundaccording to claim 26 wherein R₁ is unsubstituted naphthyl, substitutednaphthyl, unsubstituted phenyl, substituted phenyl, unsubstituted benzylor substituted benzyl.
 28. A compound according to claim 27 wherein R₁is benzyl.
 29. A compound according to claim 25 wherein R₁ is cyclohexylor cyclohexylmethyl.
 30. A compound according to claim 27 wherein Het is31. A compound according to claim 1 wherein R₃ is
 32. A compoundaccording to claim 31, wherein R₃ is and W is nitrogren.
 33. A compoundaccording to claim 31, wherein R₄ is selected from the group consistingof:(a) hydrogen, (b) alkyl of 1 to 6 carbon atoms or alkyl of 1 to 6carbon atoms substituted with Z₅, wherein Z₅ is selected from the groupconsisting of hydroxy, halogen, --C(O)OH, --C(O)OR₈, --S(O)₂ OH and--S(O)_(p) R₈, (c) alkyl of 1 to 3 carbon atoms substituted with cyclicalkyl of 3 to 5 carbon atoms, (d) alkenyl of about 3 to about 6 carbonatoms, (e) cycloalkyl of about 3 to about 5 carbon atoms, (f) heteroarylof 5 atoms, and (g) heteroaralkyl of 6 atoms.
 34. A compound accordingto claim 33, wherein R₅ is hydrogen.
 35. A compound according to claim31, wherein R₆ is selected from the group consisting of:(a) hydrogen,(b) alkyl of 1 to about 12 carbon atoms or alkyl of 1 to 12 carbon atomssubstituted with Z₆, wherein Z₆ is selected from the group consisting ofhydroxy, halogen, --OR₉, --NHR₉, --C(O)OH, --C(O)OR₉, --S(O)₂ OH and--S(O)_(p) R₉, (c) alkyl of 1 to about 3 carbon atoms substituted withcyclic alkyl of about 5 to about 8 carbon atoms, (d) alkenyl of about 2to about 6 carbon atoms which is optionally substituted with cyclicalkyl of about 3 to about 8 carbon atoms, or aryl of about 6 to about 10carbon atoms, (e) aralkyl or substituted aralkyl, (f) heteroaralkyl ofabout 5 to 10 ring atoms or substituted heteroaralkyl of about 5 to 10ring atoms, (g) aralkenyl of about 8 to 15 carbon atoms which isoptionally mono-, di- or tri-substituted on the ring with Y₁, Y₂ and/orY₃, and (h) heteroaralkenyl of about 5 to 10 ring atoms or substitutedheteroaralkenyl of about 5 to 10 ring atoms.
 36. A compound according toclaim 35, wherein R₄ and R₅ are hydrogen or methyl and R₆ is selectedfrom the group consisting of aralkyl of about 8 to about 13 carbonatoms, and --O-aralkyl, --NH-aralkyl, or --S(O)_(p) -aralkyl all ofabout 7 to about 12 carbon atoms.
 37. A compound according to claim 36,wherein the aryl portion of the aralkyl group of R₆ is selected fromunsubstituted or substituted phenyl or naphthyl.
 38. A compoundaccording to claim 37, wherein said substituents of the aryl ring areselected from the group consisting of methyl, methoxy, fluoro, chloroand trifluoromethyl.
 39. A compound according to claim 35, wherein R₆ isselected from the group consisting of phenylethyl, phenylpropyl,cyclohexylethyl and cyclohexylpropyl.
 40. A compound according to claim31, wherein R₇ is selected from the group consisting of hydrogen,methyl, difluoromethyl and trifluoromethyl.
 41. A compound according toclaim 40, wherein R₇ is hydrogen.
 42. A compound according to claim 31,wherein Het is ##STR239## wherein R₄ is selected from the groupconsisting of hydrogen, methyl, ethyl, propenyl, allyl, propyl,isopropyl, butyl, R-sec-butyl, S-sec-butyl, isobutyl, 1-pentyl,R-2-pentyl, S-2-pentyl, 3-pentyl, S-1-(2-methyl)-butyl,R-2-(3-methyl)-butyl, 1-(3-methyl)-butyl, R-1-(2-methyl)-butyl,cyclopentyl, 2-pyrolyl, 3-pyrolyl, 1-hexyl, S-2-hexyl, R-2-hexyl,R-3-hexyl, and S-3-hexyl and R₅ and R₆ are independently selected fromhydrogen and methyl.
 43. A compound according to claim 42, wherein R₄ ishydrogen or methyl.
 44. A compound according to claim 31, wherein X is--S(O)₂ --, R₁ is substituted or unsubstituted aralkyl or substituted orunsubstituted aryl, R₃ is ##STR240## wherein W is nitrogen, and Het is##STR241##
 45. A compound according to claim 44, wherein R₁ issubstituted or unsubstituted benzyl or phenyl.
 46. A compound accordingto claim 31 wherein X is --S(O)₂ --.
 47. A compound according to claim46 wherein R₁ is alkyl, aryl or aralkyl.
 48. A compound according toclaim 47 wherein R₁ is aryl or aralkyl optionally substituted with Y₁and Y₂ and Y₁ and Y₂ are independently selected from --C(O)OH,--C(O)OZ₁, --OH, --S(O)_(m) Z₁, and --CF₃.
 49. A compound according toclaim 48 wherein R₁ is unsubstituted naphthyl, substituted naphthyl,unsubstituted phenyl, substituted phenyl, unsubstituted benzyl orsubstituted benzyl.
 50. A compound according to claim 49 wherein R₁ isbenzyl.
 51. A compound according to claim 49 wherein R₁ is cyclohexyl orcyclohexylmethyl.
 52. A compound according to claim 44 wherein Het is53. A compound according to claim 32 wherein X is --S(O)₂ --.
 54. Acompound according to claim 53 wherein R₁ is alkyl, cycloalkyl, aryl, oraralkyl.
 55. A compound according to claim 54 wherein R₁ is aryl oraralkyl optionally substituted with Y₁ and Y₂ and wherein Y₁ and Y₂ areindependently selected from --C(O)OH, --C(O)OZ₁, --OH, --S(O)_(m) Z₁,and --CF₃.
 56. A compound according to claim 55 wherein R₁ isunsubstituted naphthyl, substituted naphthyl, unsubstituted phenyl,substituted phenyl, unsubstituted benzyl, or substituted benzyl.
 57. Acompound according to claim 56 wherein R₁ is benzyl.
 58. A compoundaccording to claim 54 wherein R₁ is cyclohexyl or cyclohexylmethyl. 59.A compound according to claim 1 having the formula:
 60. A compoundaccording to claim 9 selected from the group consisting of:3-(phenylsulfonyl)amino-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(2-naphthylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(1-naphthylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-(cyclohexylaminosulfonyl)amino-2-oxo-1,2-dihydropyridyl-acetyl-L-argininal,3-(phenylaminosulfonyl)amino-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(phenoxycarbonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(cyclohexylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,-[(cyclohexylmethyloulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(phenethylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-((2-methoxycarbonylphenylsulfonyl)amino)-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(3-methoxycarbonylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(4-methoxycarbonylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(2-trifluoromethylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(3-trifluoromethylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(4-trifluoromethylphenylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(2-methoxycarbonylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(3 -methoxycarbonylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(4-methoxycarbonylbenzylsulfonyl)amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(2-trifluoromethylbenzylsulfonyl) amino]-2-oxo-1,2-dihydropyridylacetyl -L-argininal, 3-[(3-trifluoromethylbenzylsulfonyl) amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal,3-[(4-trifluoromethylbenzylsulfonyl) amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal, (3-[(benzylsulfonyl)amino]-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,[3-[(benzylsulfonyl)amino]-6-methyl-2-oxo-1,2-dihydropyridyl]acetyl-L-argininal,and 3,-[(2-trifluoromethylbenzylsulfonyl) amino]-2-oxo-1,2-dihydropyridylacetyl-L-argininal.
 61. A compound according toclaim 31, wherein R₅ is selected from the group consisting of hydrogen,alkyl of 1 to about 5 carbon atoms, trifluoromethyl, and alkoxy of 1 to4 carbon atoms.
 62. A method of alkylating a3-nitro-2-oxo-1,2-dihydropyridyl acetate compound at ring position 4comprising(a) combining the compound with a solution of a zinc salt andan alkyl grignard under anhydrous conditions to form a3-nitro-2-oxo-4-alkyl-1,2,3,4-dihydropyridyl acetate intermediate, (b)contacting the intermediate with an oxidizing agent, and (c) recoveringa 4-alkyl-3-nitro-2-oxo-1,2-dihydropyridyl acetate product.
 63. Themethod of claim 62, wherein the zinc salt is zinc chloride or zincbromide.
 64. The method of claim 63, wherein the oxidizing agent ispalladium acetate in warm THF.
 65. The method of claim 62, wherein instep (a) the compound and zinc salt are first combined and then thealkyl grignard is added.
 66. The method of claim 62, wherein the alkylgrignard is synthesized from a starting compound having an R₁ of formula(I) selected from an alkyl or aryl bromide, chloride or iodide whereinsaid R₁ group is selected from the group consisting of(1) alkyl of 1 toabout 12 carbon atoms, (2) alkyl of 1 to about 3 carbon atomssubstituted with cyclic alkyl of about 3 to about 8 carbon atoms, whichoptionally is substituted in the ring carbons with hydroxyl, amino,guanidino, amidino, or alkoxyl or alkyl each of 1 to about 3 carbons,(3) cyclic alkyl of 3 to about 15 carbon atoms, which optionally issubstituted in the ring carbons with hydroxyl, amino, quanidino,amidino, or alkoxyl or alkyl each of 1 to about 3 carbons, (4) aryl ofabout 6 to about 14 carbon atoms which is optionally mono-, di-, ortri-substituted with Y₁, Y₂, and/or Y₃, and (5) aralkyl of about 7 toabout 15 carbon atoms which is optionally substituted on the alkyl chainwith hydroxy or halogen and optionally mono-, di-, or tri-substituted onthe aryl ring with Y₁, Y₂, and/or Y₃, wherein Y₁, Y₂, and Y₃ are(i)independently selected from the group consisting of hydrogen, halogen,cyano, tetrazolyl, amino, guanidino, amidino, methylamino, andmethylguanidino, --CF₃, CF₂ H, --CF₂ CF₃, --CH(CF₃) 2, --C (OH) (CF₃) 2,--OCF₃, --OCF₂ CF₃, --OC(O)NH₂, --OC(O)NHZ₁, --OC(O) NZ₁ Z₂, --NHC (O)Z₁, --NHC(O)NH₂, --NHC(O)NZ₁, --NHC(O) NZ₁ Z₂, --C (O) OH, --C(O)NH₂,--C (O) NHZ₁, --C (O) OZ₁, --P (0) ₃ H₂, --P(O)₃ (Z₁)₂, --S(O)₃ H,--S(O)_(m) Z₁, --Z₁, --OZ₁, --OH, --NH₂, --NHZ₁, and --NZ₁ Z₂, wherein mis 0, 1 or 2, and Z₁ and Z₂ are independently selected from the groupconsisting of alkyl of 1 to about 12 carbon atoms, aryl of about 6 toabout 14 carbon atoms, heteroaryl of about 5 to about 14 atoms having 1to about 9 carbon atoms, aralkyl of about 7 to about 15 carbon atoms,and heteroaralkyl of about 6 to about 11 atoms having about 3 to about 9carbon atoms, or (ii) Y₁ and Y₂ are selected together to be --OC(Z₃)(Z₄)O--, wherein Z₃ and Z₄ are independently selected from the groupconsisting of hydrogen, alkyl of 1 to about 12 carbon atoms, aryl ofabout 6 to about 14 carbon atoms, heteroaryl of about 5 to about 14atoms having 1 to about 9 carbon atoms, aralkyl of about 7 to about 15carbon atoms, and heteroaralkyl of about 6 to about 11 atoms havingabout 3 to about 9 carbon atoms.
 67. The method of claim 66, wherein thealkyl grignard is 3-phenylpropyl magnesium bromide.